Multi-imaging analysis of nascent surface structures generated during femtosecond laser irradiation of silicon in high vacuum

NASA Astrophysics Data System (ADS)

Gesuele, F.; JJ Nivas, J.; Fittipaldi, R.; Altucci, C.; Bruzzese, R.; Maddalena, P.; Amoruso, S.

2018-02-01

We report a correlative imaging analysis of a crystalline silicon target after irradiation with a low number of 1055 nm, 850 fs laser pulses with several microscopy techniques (e.g., scanning electron microscopy, atomic force microscopy, Raman micro-imaging and confocal optical microscopy). The analysis is carried out on samples irradiated both in high vacuum and at atmospheric pressure conditions, evidencing interesting differences induced by the ambient environment. In high-vacuum conditions, the results evidence the formation of a halo, which is constituted by alternate stripes of amorphous and crystalline silicon, around the nascent ablation crater. In air, such an effect is drastically reduced, due to the significant back-deposition of nanoparticulate material induced by the larger ambient pressure.

Analysis-Preserving Video Microscopy Compression via Correlation and Mathematical Morphology

PubMed Central

Shao, Chong; Zhong, Alfred; Cribb, Jeremy; Osborne, Lukas D.; O’Brien, E. Timothy; Superfine, Richard; Mayer-Patel, Ketan; Taylor, Russell M.

2015-01-01

The large amount video data produced by multi-channel, high-resolution microscopy system drives the need for a new high-performance domain-specific video compression technique. We describe a novel compression method for video microscopy data. The method is based on Pearson's correlation and mathematical morphology. The method makes use of the point-spread function (PSF) in the microscopy video acquisition phase. We compare our method to other lossless compression methods and to lossy JPEG, JPEG2000 and H.264 compression for various kinds of video microscopy data including fluorescence video and brightfield video. We find that for certain data sets, the new method compresses much better than lossless compression with no impact on analysis results. It achieved a best compressed size of 0.77% of the original size, 25× smaller than the best lossless technique (which yields 20% for the same video). The compressed size scales with the video's scientific data content. Further testing showed that existing lossy algorithms greatly impacted data analysis at similar compression sizes. PMID:26435032

Co-axial Electrospun Polyacrylonitrile-Poly(methylmethacrylate) Nanofibers: Atomic Force Microscopy and Compositional Characterization

PubMed Central

Zander, N.E.; Strawhecker, K.E.; Orlicki, J.A.; Rawlett, A.M.; Beebe, T.P.

2011-01-01

Poly(methylmethacrylate) (PMMA)- Polyacrylonitrile (PAN) fibers were prepared using a conventional single-nozzle electrospinning technique. The as-spun fibers exhibited core-shell morphology as verified by transmission electron microscopy (TEM) and atomic force microscopy (AFM). AFM-phase and modulus mapping images of the fiber cross-section and x-ray photoelectron spectroscopy (XPS) analysis indicated PAN formed the shell and PMMA the core material. XPS, thermal gravimetric analysis (TGA), and elemental analysis were used to determine fiber compositional information. Soaking the fibers in solvent demonstrated removal of the core material, generating hollow PAN fibers. PMID:21928836

Retracing in correlative light electron microscopy: where is my object of interest?

PubMed

Hodgson, Lorna; Nam, David; Mantell, Judith; Achim, Alin; Verkade, Paul

2014-01-01

Correlative light electron microscopy (CLEM) combines the strengths of light and electron microscopy in a single experiment. There are many ways to perform a CLEM experiment and a variety of microscopy modalities can be combined either on separate instruments or as completely integrated solutions. In general, however, a CLEM experiment can be divided into three parts: probes, processing, and analysis. Most of the existing technologies are focussed around the development and use of probes or describe processing methodologies that explain or circumvent some of the compromises that need to be made when performing both light and electron microscopy on the same sample. So far, relatively little attention has been paid to the analysis part of CLEM experiments. Although it is an essential part of each CLEM experiment, it is usually a cumbersome manual process. Here, we briefly discuss each of the three above-mentioned steps, with a focus on the analysis part. We will also introduce an automated registration algorithm that can be applied to the analysis stage to enable the accurate registration of LM and EM images. This facilitates tracing back the right cell/object seen in the light microscope in the EM. © 2014 Elsevier Inc. All rights reserved.

Advanced electron microscopy methods for the analysis of MgB2 superconductor

NASA Astrophysics Data System (ADS)

Birajdar, B.; Peranio, N.; Eibl, O.

2008-02-01

Advanced electron microscopy methods used for the analysis of superconducting MgB2 wires and tapes are described. The wires and tapes were prepared by the powder in tube method using different processing technologies and thoroughly characterised for their superconducting properties within the HIPERMAG project. Microstructure analysis on μm to nm length scales is necessary to understand the superconducting properties of MgB2. For the MgB2 phase analysis on μm scale an analytical SEM, and for the analysis on nm scale a energy-filtered STEM is used. Both the microscopes were equipped with EDX detector and field emission gun. Electron microscopy and spectroscopy of MgB2 is challenging because of the boron analysis, carbon and oxygen contamination, and the presence of large number of secondary phases. Advanced electron microscopy involves, combined SEM, EPMA and TEM analysis with artefact free sample preparation, elemental mapping and chemical quantification of point spectra. Details of the acquisition conditions and achieved accuracy are presented. Ex-situ wires show oxygen-free MgB2 colonies (a colony is a dense arrangement of several MgB2 grains) embedded in a porous and oxygen-rich matrix, introducing structural granularity. In comparison, in-situ wires are generally more dense, but show inhibited MgB2 phase formation with significantly higher fraction of B-rich secondary phases. SiC additives in the in-situ wires forms Mg2Si secondary phases. The advanced electron microscopy has been used to extract the microstructure parameters like colony size, B-rich secondary phase fraction, O mole fraction and MgB2 grain size, and establish a microstructure-critical current density model [1]. In summary, conventional secondary electron imaging in SEM and diffraction contrast imaging in the TEM are by far not sufficient and advanced electron microscopy methods are essential for the analysis of superconducting MgB2 wires and tapes.

Microscopy and Image Analysis.

PubMed

McNamara, George; Difilippantonio, Michael; Ried, Thomas; Bieber, Frederick R

2017-07-11

This unit provides an overview of light microscopy, including objectives, light sources, filters, film, and color photography for fluorescence microscopy and fluorescence in situ hybridization (FISH). We believe there are excellent opportunities for cytogeneticists, pathologists, and other biomedical readers, to take advantage of specimen optical clearing techniques and expansion microscopy-we briefly point to these new opportunities. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

Automated analysis of high-content microscopy data with deep learning.

PubMed

Kraus, Oren Z; Grys, Ben T; Ba, Jimmy; Chong, Yolanda; Frey, Brendan J; Boone, Charles; Andrews, Brenda J

2017-04-18

Existing computational pipelines for quantitative analysis of high-content microscopy data rely on traditional machine learning approaches that fail to accurately classify more than a single dataset without substantial tuning and training, requiring extensive analysis. Here, we demonstrate that the application of deep learning to biological image data can overcome the pitfalls associated with conventional machine learning classifiers. Using a deep convolutional neural network (DeepLoc) to analyze yeast cell images, we show improved performance over traditional approaches in the automated classification of protein subcellular localization. We also demonstrate the ability of DeepLoc to classify highly divergent image sets, including images of pheromone-arrested cells with abnormal cellular morphology, as well as images generated in different genetic backgrounds and in different laboratories. We offer an open-source implementation that enables updating DeepLoc on new microscopy datasets. This study highlights deep learning as an important tool for the expedited analysis of high-content microscopy data. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Faster tissue interface analysis from Raman microscopy images using compressed factorisation

NASA Astrophysics Data System (ADS)

Palmer, Andrew D.; Bannerman, Alistair; Grover, Liam; Styles, Iain B.

2013-06-01

The structure of an artificial ligament was examined using Raman microscopy in combination with novel data analysis. Basis approximation and compressed principal component analysis are shown to provide efficient compression of confocal Raman microscopy images, alongside powerful methods for unsupervised analysis. This scheme allows the acceleration of data mining, such as principal component analysis, as they can be performed on the compressed data representation, providing a decrease in the factorisation time of a single image from five minutes to under a second. Using this workflow the interface region between a chemically engineered ligament construct and a bone-mimic anchor was examined. Natural ligament contains a striated interface between the bone and tissue that provides improved mechanical load tolerance, a similar interface was found in the ligament construct.

Lensfree On-Chip Microscopy and Tomography for Bio-Medical Applications

PubMed Central

Isikman, Serhan O.; Bishara, Waheb; Mudanyali, Onur; Sencan, Ikbal; Su, Ting-Wei; Tseng, Derek; Yaglidere, Oguzhan; Sikora, Uzair; Ozcan, Aydogan

2012-01-01

Lensfree on-chip holographic microscopy is an emerging technique that offers imaging of biological specimens over a large field-of-view without using any lenses or bulky optical components. Lending itself to a compact, cost-effective and mechanically robust architecture, lensfree on-chip holographic microscopy can offer an alternative toolset addressing some of the emerging needs of microscopic analysis and diagnostics in low-resource settings, especially for telemedicine applications. In this review, we summarize the latest achievements in lensfree optical microscopy based on partially coherent on-chip holography, including portable telemedicine microscopy, cell-phone based microscopy and field-portable optical tomographic microscopy. We also discuss some of the future directions for telemedicine microscopy and its prospects to help combat various global health challenges. PMID:24478572

Genetically encoded sensors and fluorescence microscopy for anticancer research

NASA Astrophysics Data System (ADS)

Zagaynova, Elena V.; Shirmanova, Marina V.; Sergeeva, Tatiana F.; Klementieva, Natalia V.; Mishin, Alexander S.; Gavrina, Alena I.; Zlobovskay, Olga A.; Furman, Olga E.; Dudenkova, Varvara V.; Perelman, Gregory S.; Lukina, Maria M.; Lukyanov, Konstantin A.

2017-02-01

Early response of cancer cells to chemical compounds and chemotherapeutic drugs were studied using novel fluorescence tools and microscopy techniques. We applied confocal microscopy, two-photon fluorescence lifetime imaging microscopy and super-resolution localization-based microscopy to assess structural and functional changes in cancer cells in vitro. The dynamics of energy metabolism, intracellular pH, caspase-3 activation during staurosporine-induced apoptosis as well as actin cytoskeleton rearrangements under chemotherapy were evaluated. We have showed that new genetically encoded sensors and advanced fluorescence microscopy methods provide an efficient way for multiparameter analysis of cell activities

Scanning probe recognition microscopy investigation of tissue scaffold properties

PubMed Central

Fan, Yuan; Chen, Qian; Ayres, Virginia M; Baczewski, Andrew D; Udpa, Lalita; Kumar, Shiva

2007-01-01

Scanning probe recognition microscopy is a new scanning probe microscopy technique which enables selective scanning along individual nanofibers within a tissue scaffold. Statistically significant data for multiple properties can be collected by repetitively fine-scanning an identical region of interest. The results of a scanning probe recognition microscopy investigation of the surface roughness and elasticity of a series of tissue scaffolds are presented. Deconvolution and statistical methods were developed and used for data accuracy along curved nanofiber surfaces. Nanofiber features were also independently analyzed using transmission electron microscopy, with results that supported the scanning probe recognition microscopy-based analysis. PMID:18203431

Scanning probe recognition microscopy investigation of tissue scaffold properties.

PubMed

Fan, Yuan; Chen, Qian; Ayres, Virginia M; Baczewski, Andrew D; Udpa, Lalita; Kumar, Shiva

2007-01-01

Scanning probe recognition microscopy is a new scanning probe microscopy technique which enables selective scanning along individual nanofibers within a tissue scaffold. Statistically significant data for multiple properties can be collected by repetitively fine-scanning an identical region of interest. The results of a scanning probe recognition microscopy investigation of the surface roughness and elasticity of a series of tissue scaffolds are presented. Deconvolution and statistical methods were developed and used for data accuracy along curved nanofiber surfaces. Nanofiber features were also independently analyzed using transmission electron microscopy, with results that supported the scanning probe recognition microscopy-based analysis.

Comparison of four methods of surface roughness assessment of corneal stromal bed after lamellar cutting

PubMed Central

Jumelle, Clotilde; Hamri, Alina; Egaud, Gregory; Mauclair, Cyril; Reynaud, Stephanie; Dumas, Virginie; Pereira, Sandrine; Garcin, Thibaud; Gain, Philippe; Thuret, Gilles

2017-01-01

Corneal lamellar cutting with a blade or femtosecond laser (FSL) is commonly used during refractive surgery and corneal grafts. Surface roughness of the cutting plane influences postoperative visual acuity but is difficult to assess reliably. For the first time, we compared chromatic confocal microscopy (CCM) with scanning electron microscopy, atomic force microscopy (AFM) and focus-variation microscopy (FVM) to characterize surfaces of variable roughness after FSL cutting. The small area allowed by AFM hinders conclusive roughness analysis, especially with irregular cuts. FVM does not always differentiate between smooth and rough surfaces. Finally, CCM allows analysis of large surfaces and differentiates between surface states. PMID:29188095

Dark Field Microscopy for Analytical Laboratory Courses

ERIC Educational Resources Information Center

Augspurger, Ashley E.; Stender, Anthony S.; Marchuk, Kyle; Greenbowe, Thomas J.; Fang, Ning

2014-01-01

An innovative and inexpensive optical microscopy experiment for a quantitative analysis or an instrumental analysis chemistry course is described. The students have hands-on experience with a dark field microscope and investigate the wavelength dependence of localized surface plasmon resonance in gold and silver nanoparticles. Students also…

Incorporating Basic Optical Microscopy in the Instrumental Analysis Laboratory

ERIC Educational Resources Information Center

Flowers, Paul A.

2011-01-01

A simple and versatile approach to incorporating basic optical microscopy in the undergraduate instrumental analysis laboratory is described. Attaching a miniature CCD spectrometer to the video port of a standard compound microscope yields a visible microspectrophotometer suitable for student investigations of fundamental spectrometry concepts,…

EVALUATION OF COMPUTER-CONTROLLED SCANNING ELECTRON MICROSCOPY APPLIED TO AN AMBIENT URBAN AEROSOL SAMPLE

EPA Science Inventory

Recent interest in monitoring and speciation of particulate matter has led to increased application of scanning electron microscopy (SEM) coupled with energy-dispersive x-ray analysis (EDX) to individual particle analysis. SEM/EDX provides information on the size, shape, co...

Nonlinear dynamic phase contrast microscopy for microfluidic and microbiological applications

NASA Astrophysics Data System (ADS)

Denz, C.; Holtmann, F.; Woerdemann, M.; Oevermann, M.

2008-08-01

In live sciences, the observation and analysis of moving living cells, molecular motors or motion of micro- and nano-objects is a current field of research. At the same time, microfluidic innovations are needed for biological and medical applications on a micro- and nano-scale. Conventional microscopy techniques are reaching considerable limits with respect to these issues. A promising approach for this challenge is nonlinear dynamic phase contrast microscopy. It is an alternative full field approach that allows to detect motion as well as phase changes of living unstained micro-objects in real-time, thereby being marker free, without contact and non destructive, i.e. fully biocompatible. The generality of this system allows it to be combined with several other microscope techniques such as conventional bright field or fluorescence microscopy. In this article we will present the dynamic phase contrast technique and its applications in analysis of micro organismic dynamics, micro flow velocimetry and micro-mixing analysis.

The Use of Contact Mode Atomic Force Microscopy in Aqueous Medium for Structural Analysis of Spinach Photosynthetic Complexes

DOE PAGES

Phuthong, Witchukorn; Huang, Zubin; Wittkopp, Tyler M.; ...

2015-07-28

To investigate the dynamics of photosynthetic pigment-protein complexes in vascular plants at high resolution in an aqueous environment, membrane-protruding oxygen-evolving complexes (OECs) associated with photosystem II (PSII) on spinach ( Spinacia oleracea) grana membranes were examined using contact mode atomic force microscopy. This study represents, to our knowledge, the first use of atomic force microscopy to distinguish the putative large extrinsic loop of Photosystem II CP47 reaction center protein (CP47) from the putative oxygen-evolving enhancer proteins 1, 2, and 3 (PsbO, PsbP, and PsbQ) and large extrinsic loop of Photosystem II CP43 reaction center protein (CP43) in the PSII-OEC extrinsicmore » domains of grana membranes under conditions resulting in the disordered arrangement of PSII-OEC particles. Moreover, we observed uncharacterized membrane particles that, based on their physical characteristics and electrophoretic analysis of the polypeptides associated with the grana samples, are hypothesized to be a domain of photosystem I that protrudes from the stromal face of single thylakoid bilayers. Furthermore, our results are interpreted in the context of the results of others that were obtained using cryo-electron microscopy (and single particle analysis), negative staining and freeze-fracture electron microscopy, as well as previous atomic force microscopy studies.« less

Crystals seen on CSF microscopy in a case of suspected subarachnoid haemorrhage

PubMed Central

Weiand, Daniel; Hanning, Ian; Mouhamadou, Moussa; Wearmouth, Debbie

2015-01-01

Although crystals are rarely identified on cerebrospinal fluid (CSF) microscopy, their presence can be of significant diagnostic value. We report a case of oxalate crystals seen on CSF microscopy of a 43-year-old woman. The patient presented with headaches, nausea and vomiting. CT of the head showed a small focus of hyper-density, suspicious of haemorrhage, in the right side of the pontine cistern. CSF cell count was within the normal range. Although no organisms were seen on microscopy, copious oxalate crystals were seen. The same crystals were seen on microscopy of CSF collected in a fluoride oxalate container used for glucose analysis. A follow-up contrast-enhanced CT angiogram did not demonstrate any abnormalities. It transpired that excess CSF had been collected into a fluoride oxalate container. This had subsequently been decanted into a plain container for microbiological analysis. Correct specimen collection should be emphasised when teaching lumbar puncture technique. PMID:26139652

New Tools for Comparing Microscopy Images: Quantitative Analysis of Cell Types in Bacillus subtilis

PubMed Central

van Gestel, Jordi; Vlamakis, Hera

2014-01-01

Fluorescence microscopy is a method commonly used to examine individual differences between bacterial cells, yet many studies still lack a quantitative analysis of fluorescence microscopy data. Here we introduce some simple tools that microbiologists can use to analyze and compare their microscopy images. We show how image data can be converted to distribution data. These data can be subjected to a cluster analysis that makes it possible to objectively compare microscopy images. The distribution data can further be analyzed using distribution fitting. We illustrate our methods by scrutinizing two independently acquired data sets, each containing microscopy images of a doubly labeled Bacillus subtilis strain. For the first data set, we examined the expression of srfA and tapA, two genes which are expressed in surfactin-producing and matrix-producing cells, respectively. For the second data set, we examined the expression of eps and tapA; these genes are expressed in matrix-producing cells. We show that srfA is expressed by all cells in the population, a finding which contrasts with a previously reported bimodal distribution of srfA expression. In addition, we show that eps and tapA do not always have the same expression profiles, despite being expressed in the same cell type: both operons are expressed in cell chains, while single cells mainly express eps. These findings exemplify that the quantification and comparison of microscopy data can yield insights that otherwise would go unnoticed. PMID:25448819

New tools for comparing microscopy images: quantitative analysis of cell types in Bacillus subtilis.

PubMed

van Gestel, Jordi; Vlamakis, Hera; Kolter, Roberto

2015-02-15

Fluorescence microscopy is a method commonly used to examine individual differences between bacterial cells, yet many studies still lack a quantitative analysis of fluorescence microscopy data. Here we introduce some simple tools that microbiologists can use to analyze and compare their microscopy images. We show how image data can be converted to distribution data. These data can be subjected to a cluster analysis that makes it possible to objectively compare microscopy images. The distribution data can further be analyzed using distribution fitting. We illustrate our methods by scrutinizing two independently acquired data sets, each containing microscopy images of a doubly labeled Bacillus subtilis strain. For the first data set, we examined the expression of srfA and tapA, two genes which are expressed in surfactin-producing and matrix-producing cells, respectively. For the second data set, we examined the expression of eps and tapA; these genes are expressed in matrix-producing cells. We show that srfA is expressed by all cells in the population, a finding which contrasts with a previously reported bimodal distribution of srfA expression. In addition, we show that eps and tapA do not always have the same expression profiles, despite being expressed in the same cell type: both operons are expressed in cell chains, while single cells mainly express eps. These findings exemplify that the quantification and comparison of microscopy data can yield insights that otherwise would go unnoticed. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Cost-utility analysis of LED fluorescence microscopy in the diagnosis of pulmonary tuberculosis in Indian settings.

PubMed

Kelly, V; Sagili, K D; Satyanarayana, S; Reza, L W; Chadha, S S; Wilson, N C

2015-06-01

With support from the Stop TB Partnership's TB REACH Wave 2 Grant, diagnostic microscopy services for tuberculosis (TB) were upgraded from conventional Ziehl-Neelsen (ZN) based sputum microscopy to light emitting diode technology-based fluorescence microscopy (LED FM) in 200 high-workload microscopy centres in India as a pilot intervention. To evaluate the cost-effectiveness of LED-FM over conventional ZN microscopy to inform further scale-up. A decision-tree model was constructed to assess the cost utility of LED FM over ZN microscopy. The results were summarised using incremental cost-effectiveness ratio (ICER); one-way and probabilistic sensitivity analyses were also conducted to address uncertainty within the model. Data were analysed from 200 medical colleges in 2011 and 2012, before and after the introduction of LED microscopes. A full costing analysis was carried out from the perspective of a national TB programme. The ICER was calculated at US$14.64 per disability-adjusted life-year, with an 82% probability of being cost-effective at a willingness-to-pay threshold equivalent to Indian gross domestic product per capita. LED FM is a cost-effective intervention for detecting TB cases in India at high-workload medical college settings.

Detection, mapping, and quantification of single walled carbon nanotubes in histological specimens with photoacoustic microscopy.

PubMed

Avti, Pramod K; Hu, Song; Favazza, Christopher; Mikos, Antonios G; Jansen, John A; Shroyer, Kenneth R; Wang, Lihong V; Sitharaman, Balaji

2012-01-01

In the present study, the efficacy of multi-scale photoacoustic microscopy (PAM) was investigated to detect, map, and quantify trace amounts [nanograms (ng) to micrograms (µg)] of SWCNTs in a variety of histological tissue specimens consisting of cancer and benign tissue biopsies (histological specimens from implanted tissue engineering scaffolds). Optical-resolution (OR) and acoustic-resolution (AR)--Photoacoustic microscopy (PAM) was employed to detect, map and quantify the SWCNTs in a variety of tissue histological specimens and compared with other optical techniques (bright-field optical microscopy, Raman microscopy, near infrared (NIR) fluorescence microscopy). Both optical-resolution and acoustic-resolution PAM, allow the detection and quantification of SWCNTs in histological specimens with scalable spatial resolution and depth penetration. The noise-equivalent detection sensitivity to SWCNTs in the specimens was calculated to be as low as ∼7 pg. Image processing analysis further allowed the mapping, distribution, and quantification of the SWCNTs in the histological sections. The results demonstrate the potential of PAM as a promising imaging technique to detect, map, and quantify SWCNTs in histological specimens, and could complement the capabilities of current optical and electron microscopy techniques in the analysis of histological specimens containing SWCNTs.

Diagnostic accuracy of same-day microscopy versus standard microscopy for pulmonary tuberculosis: a systematic review and meta-analysis.

PubMed

Davis, J Lucian; Cattamanchi, Adithya; Cuevas, Luis E; Hopewell, Philip C; Steingart, Karen R

2013-02-01

Sputum smear microscopy is the most widely available diagnostic test for pulmonary tuberculosis in countries with a high burden of the disease. Improving its accuracy is crucial to achievement of case-detection targets established by the Millennium Development Goals. Unfortunately, many patients are unable to submit all of the specimens needed for examination or to return for treatment because standard sputum collection and reporting requires several clinic visits. To inform policy recommendations by a WHO-convened Expert Group, we aimed to assess the accuracy of sputum smear examination with strategies for obtaining sputum on 1 day compared with strategies for obtaining sputum over 2 days. We did a systematic review and meta-analysis of research articles comparing the accuracy of front-loaded or same-day microscopy and standard sputum smear microscopy for diagnosis of culture-confirmed pulmonary tuberculosis. We searched Medline, Embase, Biosis, and Web of Science for articles published between Jan 1, 2005, and Feb 14, 2012. Two investigators identified eligible articles and extracted data for individual study sites. We generated pooled summary estimates (95% CIs) for sensitivity and specificity by use of random-effects meta-analysis when four or more studies were available. We identified eight relevant studies from five articles enrolling 7771 patients with suspected tuberculosis in low-income countries. Compared with the standard approach of examination of two smears with Ziehl-Neelsen light microscopy over 2 days, examination of two smears taken on the same day had much the same sensitivity (64% [95% CI 60 to 69] for standard microscopy vs 63% [58 to 68] for same-day microscopy) and specificity (98% [97 to 99] vs 98% [97 to 99]). We noted similar results for studies employing light-emitting diode fluorescence microscopy and for studies examining three smears, whether they were compared with two-smear strategies or with one another. Same-day sputum smear microscopy is as accurate as standard smear microscopy. Data from tuberculosis programmes are needed to document the changes required in the health system to successfully implement the strategy and understand its effects. WHO and US National Institutes of Health. Copyright © 2013 Elsevier Ltd. All rights reserved.

Spectral mapping tools from the earth sciences applied to spectral microscopy data.

PubMed

Harris, A Thomas

2006-08-01

Spectral imaging, originating from the field of earth remote sensing, is a powerful tool that is being increasingly used in a wide variety of applications for material identification. Several workers have used techniques like linear spectral unmixing (LSU) to discriminate materials in images derived from spectral microscopy. However, many spectral analysis algorithms rely on assumptions that are often violated in microscopy applications. This study explores algorithms originally developed as improvements on early earth imaging techniques that can be easily translated for use with spectral microscopy. To best demonstrate the application of earth remote sensing spectral analysis tools to spectral microscopy data, earth imaging software was used to analyze data acquired with a Leica confocal microscope with mechanical spectral scanning. For this study, spectral training signatures (often referred to as endmembers) were selected with the ENVI (ITT Visual Information Solutions, Boulder, CO) "spectral hourglass" processing flow, a series of tools that use the spectrally over-determined nature of hyperspectral data to find the most spectrally pure (or spectrally unique) pixels within the data set. This set of endmember signatures was then used in the full range of mapping algorithms available in ENVI to determine locations, and in some cases subpixel abundances of endmembers. Mapping and abundance images showed a broad agreement between the spectral analysis algorithms, supported through visual assessment of output classification images and through statistical analysis of the distribution of pixels within each endmember class. The powerful spectral analysis algorithms available in COTS software, the result of decades of research in earth imaging, are easily translated to new sources of spectral data. Although the scale between earth imagery and spectral microscopy is radically different, the problem is the same: mapping material locations and abundances based on unique spectral signatures. (c) 2006 International Society for Analytical Cytology.

Automatic segmentation of time-lapse microscopy images depicting a live Dharma embryo.

PubMed

Zacharia, Eleni; Bondesson, Maria; Riu, Anne; Ducharme, Nicole A; Gustafsson, Jan-Åke; Kakadiaris, Ioannis A

2011-01-01

Biological inferences about the toxicity of chemicals reached during experiments on the zebrafish Dharma embryo can be greatly affected by the analysis of the time-lapse microscopy images depicting the embryo. Among the stages of image analysis, automatic and accurate segmentation of the Dharma embryo is the most crucial and challenging. In this paper, an accurate and automatic segmentation approach for the segmentation of the Dharma embryo data obtained by fluorescent time-lapse microscopy is proposed. Experiments performed in four stacks of 3D images over time have shown promising results.

Biostatistical analysis of quantitative immunofluorescence microscopy images.

PubMed

Giles, C; Albrecht, M A; Lam, V; Takechi, R; Mamo, J C

2016-12-01

Semiquantitative immunofluorescence microscopy has become a key methodology in biomedical research. Typical statistical workflows are considered in the context of avoiding pseudo-replication and marginalising experimental error. However, immunofluorescence microscopy naturally generates hierarchically structured data that can be leveraged to improve statistical power and enrich biological interpretation. Herein, we describe a robust distribution fitting procedure and compare several statistical tests, outlining their potential advantages/disadvantages in the context of biological interpretation. Further, we describe tractable procedures for power analysis that incorporates the underlying distribution, sample size and number of images captured per sample. The procedures outlined have significant potential for increasing understanding of biological processes and decreasing both ethical and financial burden through experimental optimization. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

CARS microscopy of Alzheimer's diseased brain tissue

NASA Astrophysics Data System (ADS)

Enejder, Annika; Kiskis, Juris; Fink, Helen; Nyberg, Lena; Thyr, Jakob; Li, Jia-Yi

2014-02-01

Alzheimer's disease (AD) is a progressive neurodegenerative disorder currently without cure, characterized by the presence of extracellular plaques surrounded by dystrophic neurites. In an effort to understand the underlying mechanisms, biochemical analysis (protein immunoblot) of plaque extracts reveals that they consist of amyloid-beta (Aβ) peptides assembled as oligomers, protofibrils and aggregates. Their spatial distribution has been confirmed by Thioflavin-S or immuno-staining with fluorescence microscopy. However, it is increasingly understood that the protein aggregation is only one of several mechanism that causes neuronal dysfunction and death. This raises the need for a more complete biochemical analysis. In this study, we have complemented 2-photon fluorescence microscopy of Thioflavin-S and Aβ immuno-stained human AD plaques with CARS microscopy. We show that the chemical build-up of AD plaques is more complex and that Aβ staining does not provide the complete picture of the spatial distribution or the molecular composition of AD plaques. CARS images provide important complementary information to that obtained by fluorescence microscopy, motivating a broader introduction of CARS microscopy in the AD research field.

Scanning electron microscopy combined with image processing technique: Analysis of microstructure, texture and tenderness in Semitendinous and Gluteus Medius bovine muscles.

PubMed

Pieniazek, Facundo; Messina, Valeria

2016-11-01

In this study the effect of freeze drying on the microstructure, texture, and tenderness of Semitendinous and Gluteus Medius bovine muscles were analyzed applying Scanning Electron Microscopy combined with image analysis. Samples were analyzed by Scanning Electron Microscopy at different magnifications (250, 500, and 1,000×). Texture parameters were analyzed by Texture analyzer and by image analysis. Tenderness by Warner-Bratzler shear force. Significant differences (p < 0.05) were obtained for image and instrumental texture features. A linear trend with a linear correlation was applied for instrumental and image features. Image texture features calculated from Gray Level Co-occurrence Matrix (homogeneity, contrast, entropy, correlation and energy) at 1,000× in both muscles had high correlations with instrumental features (chewiness, hardness, cohesiveness, and springiness). Tenderness showed a positive correlation in both muscles with image features (energy and homogeneity). Combing Scanning Electron Microscopy with image analysis can be a useful tool to analyze quality parameters in meat.Summary SCANNING 38:727-734, 2016. © 2016 Wiley Periodicals, Inc. © Wiley Periodicals, Inc.

Comparative analysis of imaging configurations and objectives for Fourier microscopy.

PubMed

Kurvits, Jonathan A; Jiang, Mingming; Zia, Rashid

2015-11-01

Fourier microscopy is becoming an increasingly important tool for the analysis of optical nanostructures and quantum emitters. However, achieving quantitative Fourier space measurements requires a thorough understanding of the impact of aberrations introduced by optical microscopes that have been optimized for conventional real-space imaging. Here we present a detailed framework for analyzing the performance of microscope objectives for several common Fourier imaging configurations. To this end, we model objectives from Nikon, Olympus, and Zeiss using parameters that were inferred from patent literature and confirmed, where possible, by physical disassembly. We then examine the aberrations most relevant to Fourier microscopy, including the alignment tolerances of apodization factors for different objective classes, the effect of magnification on the modulation transfer function, and vignetting-induced reductions of the effective numerical aperture for wide-field measurements. Based on this analysis, we identify an optimal objective class and imaging configuration for Fourier microscopy. In addition, the Zemax files for the objectives and setups used in this analysis have been made publicly available as a resource for future studies.

Comparison of pre-processing techniques for fluorescence microscopy images of cells labeled for actin.

PubMed

Muralidhar, Gautam S; Channappayya, Sumohana S; Slater, John H; Blinka, Ellen M; Bovik, Alan C; Frey, Wolfgang; Markey, Mia K

2008-11-06

Automated analysis of fluorescence microscopy images of endothelial cells labeled for actin is important for quantifying changes in the actin cytoskeleton. The current manual approach is laborious and inefficient. The goal of our work is to develop automated image analysis methods, thereby increasing cell analysis throughput. In this study, we present preliminary results on comparing different algorithms for cell segmentation and image denoising.

Lipid Vesicle Shape Analysis from Populations Using Light Video Microscopy and Computer Vision

PubMed Central

Zupanc, Jernej; Drašler, Barbara; Boljte, Sabina; Kralj-Iglič, Veronika; Iglič, Aleš; Erdogmus, Deniz; Drobne, Damjana

2014-01-01

We present a method for giant lipid vesicle shape analysis that combines manually guided large-scale video microscopy and computer vision algorithms to enable analyzing vesicle populations. The method retains the benefits of light microscopy and enables non-destructive analysis of vesicles from suspensions containing up to several thousands of lipid vesicles (1–50 µm in diameter). For each sample, image analysis was employed to extract data on vesicle quantity and size distributions of their projected diameters and isoperimetric quotients (measure of contour roundness). This process enables a comparison of samples from the same population over time, or the comparison of a treated population to a control. Although vesicles in suspensions are heterogeneous in sizes and shapes and have distinctively non-homogeneous distribution throughout the suspension, this method allows for the capture and analysis of repeatable vesicle samples that are representative of the population inspected. PMID:25426933

Nanoscale deformation analysis with high-resolution transmission electron microscopy and digital image correlation

DOE PAGES

Wang, Xueju; Pan, Zhipeng; Fan, Feifei; ...

2015-09-10

We present an application of the digital image correlation (DIC) method to high-resolution transmission electron microscopy (HRTEM) images for nanoscale deformation analysis. The combination of DIC and HRTEM offers both the ultrahigh spatial resolution and high displacement detection sensitivity that are not possible with other microscope-based DIC techniques. We demonstrate the accuracy and utility of the HRTEM-DIC technique through displacement and strain analysis on amorphous silicon. Two types of error sources resulting from the transmission electron microscopy (TEM) image noise and electromagnetic-lens distortions are quantitatively investigated via rigid-body translation experiments. The local and global DIC approaches are applied for themore » analysis of diffusion- and reaction-induced deformation fields in electrochemically lithiated amorphous silicon. As a result, the DIC technique coupled with HRTEM provides a new avenue for the deformation analysis of materials at the nanometer length scales.« less

Changes of the elemental distributions in marine diatoms as a reporter of sample preparation artefacts. A nuclear microscopy application

NASA Astrophysics Data System (ADS)

Godinho, R. M.; Cabrita, M. T.; Alves, L. C.; Pinheiro, T.

2015-04-01

Studies of the elemental composition of whole marine diatoms cells have high interest as they constitute a direct measurement of environmental changes, and allow anticipating consequences of anthropogenic alterations to organisms, ecosystems and global marine geochemical cycles. Nuclear microscopy is a powerful tool allowing direct measurement of whole cells giving qualitative imaging of distribution, and quantitative determination of intracellular concentration. Major obstacles to the analysis of marine microalgae are high medium salinity and the recurrent presence of extracellular exudates produced by algae to maintain colonies in natural media and in vitro. The objective of this paper was to optimize the methodology of sample preparation of marine unicellular algae for elemental analysis with nuclear microscopy, allowing further studies on cellular response to metals. Primary cultures of Coscinodiscus wailesii maintained in vitro were used to optimize protocols for elemental analysis with nuclear microscopy techniques. Adequate cell preparation procedures to isolate the cells from media components and exudates were established. The use of chemical agents proved to be inappropriate for elemental determination and for intracellular morphological analysis. The assessment of morphology and elemental partitioning in cell compartments obtained with nuclear microscopy techniques enabled to infer their function in natural environment and imbalances in exposure condition. Exposure to metal affected C. wailesii morphology and internal elemental distribution.

Learning a cost function for microscope image segmentation.

PubMed

Nilufar, Sharmin; Perkins, Theodore J

2014-01-01

Quantitative analysis of microscopy images is increasingly important in clinical researchers' efforts to unravel the cellular and molecular determinants of disease, and for pathological analysis of tissue samples. Yet, manual segmentation and measurement of cells or other features in images remains the norm in many fields. We report on a new system that aims for robust and accurate semi-automated analysis of microscope images. A user interactively outlines one or more examples of a target object in a training image. We then learn a cost function for detecting more objects of the same type, either in the same or different images. The cost function is incorporated into an active contour model, which can efficiently determine optimal boundaries by dynamic programming. We validate our approach and compare it to some standard alternatives on three different types of microscopic images: light microscopy of blood cells, light microscopy of muscle tissue sections, and electron microscopy cross-sections of axons and their myelin sheaths.

Microchemical Analysis Of Space Operation Debris

NASA Technical Reports Server (NTRS)

Cummings, Virginia J.; Kim, Hae Soo

1995-01-01

Report discusses techniques used in analyzing debris relative to space shuttle operations. Debris collected from space shuttle, expendable launch vehicles, payloads carried by space shuttle, and payloads carried by expendable launch vehicles. Optical microscopy, scanning electron microscopy with energy-dispersive spectrometry, analytical electron microscopy with wavelength-dispersive spectrometry, and X-ray diffraction chosen as techniques used in examining samples of debris.

Detection, Mapping, and Quantification of Single Walled Carbon Nanotubes in Histological Specimens with Photoacoustic Microscopy

PubMed Central

Mikos, Antonios G.; Jansen, John A.; Shroyer, Kenneth R.; Wang, Lihong V.; Sitharaman, Balaji

2012-01-01

Aims In the present study, the efficacy of multi-scale photoacoustic microscopy (PAM) was investigated to detect, map, and quantify trace amounts [nanograms (ng) to micrograms (µg)] of SWCNTs in a variety of histological tissue specimens consisting of cancer and benign tissue biopsies (histological specimens from implanted tissue engineering scaffolds). Materials and Methods Optical-resolution (OR) and acoustic-resolution (AR) - Photoacoustic microscopy (PAM) was employed to detect, map and quantify the SWCNTs in a variety of tissue histological specimens and compared with other optical techniques (bright-field optical microscopy, Raman microscopy, near infrared (NIR) fluorescence microscopy). Results Both optical-resolution and acoustic-resolution PAM, allow the detection and quantification of SWCNTs in histological specimens with scalable spatial resolution and depth penetration. The noise-equivalent detection sensitivity to SWCNTs in the specimens was calculated to be as low as ∼7 pg. Image processing analysis further allowed the mapping, distribution, and quantification of the SWCNTs in the histological sections. Conclusions The results demonstrate the potential of PAM as a promising imaging technique to detect, map, and quantify SWCNTs in histological specimens, and could complement the capabilities of current optical and electron microscopy techniques in the analysis of histological specimens containing SWCNTs. PMID:22496892

Contributed review: Review of integrated correlative light and electron microscopy.

PubMed

Timmermans, F J; Otto, C

2015-01-01

New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemically or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.

Extracting microtubule networks from superresolution single-molecule localization microscopy data

PubMed Central

Zhang, Zhen; Nishimura, Yukako; Kanchanawong, Pakorn

2017-01-01

Microtubule filaments form ubiquitous networks that specify spatial organization in cells. However, quantitative analysis of microtubule networks is hampered by their complex architecture, limiting insights into the interplay between their organization and cellular functions. Although superresolution microscopy has greatly facilitated high-resolution imaging of microtubule filaments, extraction of complete filament networks from such data sets is challenging. Here we describe a computational tool for automated retrieval of microtubule filaments from single-molecule-localization–based superresolution microscopy images. We present a user-friendly, graphically interfaced implementation and a quantitative analysis of microtubule network architecture phenotypes in fibroblasts. PMID:27852898

Quantitative analysis on collagen of dermatofibrosarcoma protuberans skin by second harmonic generation microscopy.

PubMed

Wu, Shulian; Huang, Yudian; Li, Hui; Wang, Yunxia; Zhang, Xiaoman

2015-01-01

Dermatofibrosarcoma protuberans (DFSP) is a skin cancer usually mistaken as other benign tumors. Abnormal DFSP resection results in tumor recurrence. Quantitative characterization of collagen alteration on the skin tumor is essential for developing a diagnostic technique. In this study, second harmonic generation (SHG) microscopy was performed to obtain images of the human DFSP skin and normal skin. Subsequently, structure and texture analysis methods were applied to determine the differences in skin texture characteristics between the two skin types, and the link between collagen alteration and tumor was established. Results suggest that combining SHG microscopy and texture analysis methods is a feasible and effective method to describe the characteristics of skin tumor like DFSP. © Wiley Periodicals, Inc.

Fluorescence Imaging of Posterior Spiracles from Second and Third Instars of Forensically-important Chrysomya rufifacies (Diptera: Calliphoridae)*

PubMed Central

Flores, Danielle; Miller, Amy L.; Showman, Angelique; Tobita, Caitlyn; Shimoda, Lori M.N.; Sung, Carl; Stokes, Alexander J.; Tomberlin, Jeffrey K.; Carter, David O.; Turner, Helen

2016-01-01

Entomological protocols for aging blow fly (Diptera: Calliphoridae) larvae to estimate the time of colonization (TOC) are commonly used to assist in death investigations. While the methodologies for analysing fly larvae differ, most rely on light microscopy, genetic analysis or, more rarely, electron microscopy. This pilot study sought to improve resolution of larval stage in the forensically-important blow fly Chrysomya rufifacies using high-content fluorescence microscopy and biochemical measures of developmental marker proteins. We established fixation and mounting protocols, defined a set of measurable morphometric criteria and captured developmental transitions of 2nd instar to 3rd instar using both fluorescence microscopy and anti-ecdysone receptor Western blot analysis. The data show that these instars can be distinguished on the basis of robust, non-bleaching, autofluorescence of larval posterior spiracles. High content imaging techniques using confocal microscopy, combined with morphometric and biochemical techniques, may therefore aid forensic entomologists in estimating TOC. PMID:27706817

Light Microscopy at Maximal Precision

NASA Astrophysics Data System (ADS)

Bierbaum, Matthew; Leahy, Brian D.; Alemi, Alexander A.; Cohen, Itai; Sethna, James P.

2017-10-01

Microscopy is the workhorse of the physical and life sciences, producing crisp images of everything from atoms to cells well beyond the capabilities of the human eye. However, the analysis of these images is frequently little more accurate than manual marking. Here, we revolutionize the analysis of microscopy images, extracting all the useful information theoretically contained in a complex microscope image. Using a generic, methodological approach, we extract the information by fitting experimental images with a detailed optical model of the microscope, a method we call parameter extraction from reconstructing images (PERI). As a proof of principle, we demonstrate this approach with a confocal image of colloidal spheres, improving measurements of particle positions and radii by 10-100 times over current methods and attaining the maximum possible accuracy. With this unprecedented accuracy, we measure nanometer-scale colloidal interactions in dense suspensions solely with light microscopy, a previously impossible feat. Our approach is generic and applicable to imaging methods from brightfield to electron microscopy, where we expect accuracies of 1 nm and 0.1 pm, respectively.

An overview of state-of-the-art image restoration in electron microscopy.

PubMed

Roels, J; Aelterman, J; Luong, H Q; Lippens, S; Pižurica, A; Saeys, Y; Philips, W

2018-06-08

In Life Science research, electron microscopy (EM) is an essential tool for morphological analysis at the subcellular level as it allows for visualization at nanometer resolution. However, electron micrographs contain image degradations such as noise and blur caused by electromagnetic interference, electron counting errors, magnetic lens imperfections, electron diffraction, etc. These imperfections in raw image quality are inevitable and hamper subsequent image analysis and visualization. In an effort to mitigate these artefacts, many electron microscopy image restoration algorithms have been proposed in the last years. Most of these methods rely on generic assumptions on the image or degradations and are therefore outperformed by advanced methods that are based on more accurate models. Ideally, a method will accurately model the specific degradations that fit the physical acquisition settings. In this overview paper, we discuss different electron microscopy image degradation solutions and demonstrate that dedicated artefact regularisation results in higher quality restoration and is applicable through recently developed probabilistic methods. © 2018 The Authors Journal of Microscopy © 2018 Royal Microscopical Society.

Introduction to Modern Methods in Light Microscopy.

PubMed

Ryan, Joel; Gerhold, Abby R; Boudreau, Vincent; Smith, Lydia; Maddox, Paul S

2017-01-01

For centuries, light microscopy has been a key method in biological research, from the early work of Robert Hooke describing biological organisms as cells, to the latest in live-cell and single-molecule systems. Here, we introduce some of the key concepts related to the development and implementation of modern microscopy techniques. We briefly discuss the basics of optics in the microscope, super-resolution imaging, quantitative image analysis, live-cell imaging, and provide an outlook on active research areas pertaining to light microscopy.

Oufti: An integrated software package for high-accuracy, high-throughput quantitative microscopy analysis

PubMed Central

Paintdakhi, Ahmad; Parry, Bradley; Campos, Manuel; Irnov, Irnov; Elf, Johan; Surovtsev, Ivan; Jacobs-Wagner, Christine

2016-01-01

Summary With the realization that bacteria display phenotypic variability among cells and exhibit complex subcellular organization critical for cellular function and behavior, microscopy has re-emerged as a primary tool in bacterial research during the last decade. However, the bottleneck in today’s single-cell studies is quantitative image analysis of cells and fluorescent signals. Here, we address current limitations through the development of Oufti, a stand-alone, open-source software package for automated measurements of microbial cells and fluorescence signals from microscopy images. Oufti provides computational solutions for tracking touching cells in confluent samples, handles various cell morphologies, offers algorithms for quantitative analysis of both diffraction and non-diffraction-limited fluorescence signals, and is scalable for high-throughput analysis of massive datasets, all with subpixel precision. All functionalities are integrated in a single package. The graphical user interface, which includes interactive modules for segmentation, image analysis, and post-processing analysis, makes the software broadly accessible to users irrespective of their computational skills. PMID:26538279

Serological testing versus other strategies for diagnosis of active tuberculosis in India: a cost-effectiveness analysis.

PubMed

Dowdy, David W; Steingart, Karen R; Pai, Madhukar

2011-08-01

Undiagnosed and misdiagnosed tuberculosis (TB) drives the epidemic in India. Serological (antibody detection) TB tests are not recommended by any agency, but widely used in many countries, including the Indian private sector. The cost and impact of using serology compared with other diagnostic techniques is unknown. Taking a patient cohort conservatively equal to the annual number of serological tests done in India (1.5 million adults suspected of having active TB), we used decision analysis to estimate costs and effectiveness of sputum smear microscopy (US$3.62 for two smears), microscopy plus automated liquid culture (mycobacterium growth indicator tube [MGIT], US$20/test), and serological testing (anda-tb ELISA, US$20/test). Data on test accuracy and costs were obtained from published literature. We adopted the perspective of the Indian TB control sector and an analysis frame of 1 year. Our primary outcome was the incremental cost per disability-adjusted life year (DALY) averted. We performed one-way sensitivity analysis on all model parameters, with multiway sensitivity analysis on variables to which the model was most sensitive. If used instead of sputum microscopy, serology generated an estimated 14,000 more TB diagnoses, but also 121,000 more false-positive diagnoses, 102,000 fewer DALYs averted, and 32,000 more secondary TB cases than microscopy, at approximately four times the incremental cost (US$47.5 million versus US$11.9 million). When added to high-quality sputum smears, MGIT culture was estimated to avert 130,000 incremental DALYs at an incremental cost of US$213 per DALY averted. Serology was dominated by (i.e., more costly and less effective than) MGIT culture and remained less economically favorable than sputum smear or TB culture in one-way and multiway sensitivity analyses. In India, sputum smear microscopy remains the most cost-effective diagnostic test available for active TB; efforts to increase access to quality-assured microscopy should take priority. In areas where high-quality microscopy exists and resources are sufficient, MGIT culture is more cost-effective than serology as an additional diagnostic test for TB. These data informed a recently published World Health Organization policy statement against serological tests.

Coherent Raman Scattering Microscopy in Biology and Medicine.

PubMed

Zhang, Chi; Zhang, Delong; Cheng, Ji-Xin

2015-01-01

Advancements in coherent Raman scattering (CRS) microscopy have enabled label-free visualization and analysis of functional, endogenous biomolecules in living systems. When compared with spontaneous Raman microscopy, a key advantage of CRS microscopy is the dramatic improvement in imaging speed, which gives rise to real-time vibrational imaging of live biological samples. Using molecular vibrational signatures, recently developed hyperspectral CRS microscopy has improved the readout of chemical information available from CRS images. In this article, we review recent achievements in CRS microscopy, focusing on the theory of the CRS signal-to-noise ratio, imaging speed, technical developments, and applications of CRS imaging in bioscience and clinical settings. In addition, we present possible future directions that the use of this technology may take.

Coherent Raman Scattering Microscopy in Biology and Medicine

PubMed Central

Zhang, Chi; Zhang, Delong; Cheng, Ji-Xin

2016-01-01

Advancements in coherent Raman scattering (CRS) microscopy have enabled label-free visualization and analysis of functional, endogenous biomolecules in living systems. When compared with spontaneous Raman microscopy, a key advantage of CRS microscopy is the dramatic improvement in imaging speed, which gives rise to real-time vibrational imaging of live biological samples. Using molecular vibrational signatures, recently developed hyperspectral CRS microscopy has improved the readout of chemical information available from CRS images. In this article, we review recent achievements in CRS microscopy, focusing on the theory of the CRS signal-to-noise ratio, imaging speed, technical developments, and applications of CRS imaging in bioscience and clinical settings. In addition, we present possible future directions that the use of this technology may take. PMID:26514285

Effects of the change in cutoff values for human epidermal growth factor receptor 2 status by immunohistochemistry and fluorescence in situ hybridization: a study comparing conventional brightfield microscopy, image analysis-assisted microscopy, and interobserver variation.

PubMed

Atkinson, Roscoe; Mollerup, Jens; Laenkholm, Anne-Vibeke; Verardo, Mark; Hawes, Debra; Commins, Deborah; Engvad, Birte; Correa, Adrian; Ehlers, Charlotte Cort; Nielsen, Kirsten Vang

2011-08-01

New guidelines for HER2 testing have been introduced. To evaluate the difference in HER2 assessment after introduction of new cutoff levels for both immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) and to compare interobserver agreement and time to score between image analysis and conventional microscopy. Samples from 150 patients with breast cancer were scored by 7 pathologists using conventional microscopy, with a cutoff of both 10% and 30% IHC-stained cells, and using automated microscopy with image analysis. The IHC results were compared individually and to HER2 status as determined by FISH, using both the approved cutoff of 2.0 and the recently introduced cutoff of 2.2. High concordance was found in IHC scoring among the 7 pathologists. The 30% cutoff led to slightly fewer positive IHC observations. Introduction of a FISH equivocal zone affected 4% of the FISH scores. If cutoff for FISH is kept at 2.0, no difference in patient selection is found between the 10% and the 30% IHC cutoff. Among the 150 breast cancer samples, the new 30% IHC and 2.2 FISH cutoff levels resulted in one case without a firm diagnosis because both IHC and FISH were equivocal. Automated microscopy and image analysis-assisted IHC led to significantly better interobserver agreement among the 7 pathologists, with an increase in mean scoring time of only about 30 seconds per slide. The change in cutoff levels led to a higher concordance between IHC and FISH, but fewer samples were classified as HER2 positive.

The potential role of in vivo reflectance confocal microscopy for evaluating oral cavity lesions: a systematic review.

PubMed

Lucchese, Alberta; Gentile, Enrica; Romano, Antonio; Maio, Claudio; Laino, Luigi; Serpico, Rosario

2016-11-01

Since the early 2000s, several studies have examined the application of reflectance confocal microscopy (RCM) to the oral cavity. This review gives an overview of the literature on reflectance confocal microscopy analysis of the oral cavity in vivo and identifies flaws in the studies, providing guidance to improve reflectance confocal microscopy applications and inform the design of future studies. The PubMed, ISI, Scopus, and Cochrane Library databases were searched for publications on RCM using the terms 'reflectance confocal microscopy' in combination with 'mouth' and other terms related to the topic of interest. The search gave 617 results. Seventeen studies were included in our final analysis. We decided to organize the selected articles according to four topics: healthy mucosa, autoimmune diseases, cancer and precancerous lesions, and hard dental tissues. Although reflectance confocal microscopy is promising for diagnosing and monitoring oral pathology, it has shortcomings and there are still too few publications on this topic. Further studies are needed to increase the quantity and quality of the results, to translate research into clinical practice. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Extent of agreement between the body fluid model of Sysmex XN-20 and the manual microscopy method.

PubMed

Huang, Wei-Hua; Lu, Lin-Peng; Wu, Kang; Guo, Fang-Yu; Guo, Jie; Yu, Jing-Long; Zhou, Dao-Yin; Sun, Yi; Deng, An-Mei

2017-09-01

Although the correlations concerning cellular component analysis between the Sysmex XN-20 body fluid (BF) model and manual microscopy have been investigated by several studies, the extent of agreement between these two methods has not been investigated. A total of 90 BF samples were prospectively collected and analyzed using the Sysmex XN-20 BF model and microscopy. The extent of agreement between these two methods was evaluated using the Bland-Altman approach. Receiver operating characteristic (ROC) curve analysis was employed to evaluate the diagnostic accuracy of high-fluorescence (HF) BF cells for malignant diseases. The agreements of white blood cell (WBC), red blood cell (RBC), and percentages of neutrophils, lymphocytes, and monocytes between the Sysmex XN-20 BF model and manual microscopy were imperfect. The areas under the ROC curves for absolute and relative HF cells were 0.67 (95% confidence interval [CI]: 0.56-0.78) and 0.60 (95% CI: 0.48-0.72), respectively. Due to the Sysmex XN-20 BF model's imperfect agreement with manual microscopy and its weak diagnostic accuracy for malignant diseases, the current evidence does not support replacing manual microscopy with this model in clinical practice. © 2016 Wiley Periodicals, Inc.

Automated urinalysis: first experiences and a comparison between the Iris iQ200 urine microscopy system, the Sysmex UF-100 flow cytometer and manual microscopic particle counting.

PubMed

Shayanfar, Noushin; Tobler, Ulrich; von Eckardstein, Arnold; Bestmann, Lukas

2007-01-01

Automated analysis of insoluble urine components can reduce the workload of conventional microscopic examination of urine sediment and is possibly helpful for standardization. We compared the diagnostic performance of two automated urine sediment analyzers and combined dipstick/automated urine analysis with that of the traditional dipstick/microscopy algorithm. A total of 332 specimens were collected and analyzed for insoluble urine components by microscopy and automated analyzers, namely the Iris iQ200 (Iris Diagnostics) and the UF-100 flow cytometer (Sysmex). The coefficients of variation for day-to-day quality control of the iQ200 and UF-100 analyzers were 6.5% and 5.5%, respectively, for red blood cells. We reached accuracy ranging from 68% (bacteria) to 97% (yeast) for the iQ200 and from 42% (bacteria) to 93% (yeast) for the UF-100. The combination of dipstick and automated urine sediment analysis increased the sensitivity of screening to approximately 98%. We conclude that automated urine sediment analysis is sufficiently precise and improves the workflow in a routine laboratory. In addition, it allows sediment analysis of all urine samples and thereby helps to detect pathological samples that would have been missed in the conventional two-step procedure according to the European guidelines. Although it is not a substitute for microscopic sediment examination, it can, when combined with dipstick testing, reduce the number of specimens submitted to microscopy. Visual microscopy is still required for some samples, namely, dysmorphic erythrocytes, yeasts, Trichomonas, oval fat bodies, differentiation of casts and certain crystals.

Electroperturbation of human stratum corneum fine structure by high voltage pulses: a freeze-fracture electron microscopy and differential thermal analysis study.

PubMed

Jadoul, A; Tanojo, H; Préat, V; Bouwstra, J A; Spies, F; Boddé, H E

1998-08-01

Application of high voltage pulses (HVP) to the skin has been shown to promote the transdermal drug delivery by a mechanism involving skin electroporation. The aim of this study was to detect potential changes in lipid phase and ultrastructure induced in human stratum corneum by various HVP protocols, using differential thermal analysis and freeze-fracture electron microscopy. Due to the time involved between the moment the electric field is switched off and the analysis, only "secondary" phenomena rather than primary events could be observed. A decrease in enthalpies for the phase transitions observed at 70 degrees C and 85 degrees C was detected by differential thermal analysis after HVP treatment. No changes in transition temperature could be seen. The freeze-fracture electron microscopy study revealed a dramatic perturbation of the lamellar ordering of the intercellular lipid after application of HVP. Most of the planes displayed rough surfaces. The lipid lamellae exhibited rounded off steps or a vanished stepwise order. There was no evidence for perturbation of the corneocytes content. In conclusion, the freeze-fracture electron microscopy and differential thermal analysis studies suggest that HVP application induces a general perturbation of the stratum corneum lipid ultrastructure.

LASER BIOLOGY: Peculiarities of studying an isolated neuron by the method of laser interference microscopy

NASA Astrophysics Data System (ADS)

Yusipovich, Alexander I.; Novikov, Sergey M.; Kazakova, Tatiana A.; Erokhova, Liudmila A.; Brazhe, Nadezda A.; Lazarev, Grigory L.; Maksimov, Georgy V.

2006-09-01

Actual aspects of using a new method of laser interference microscopy (LIM) for studying nerve cells are discussed. The peculiarities of the LIM display of neurons are demonstrated by the example of isolated neurons of a pond snail Lymnaea stagnalis. A comparative analysis of the images of the cell and subcellular structures of a neuron obtained by the methods of interference microscopy, optical transmission microscopy, and confocal microscopy is performed. Various aspects of the application of LIM for studying the lateral dimensions and internal structure of the cytoplasm and organelles of a neuron in cytology and cell physiology are discussed.

Single-molecule fluorescence microscopy review: shedding new light on old problems

PubMed Central

Shashkova, Sviatlana

2017-01-01

Fluorescence microscopy is an invaluable tool in the biosciences, a genuine workhorse technique offering exceptional contrast in conjunction with high specificity of labelling with relatively minimal perturbation to biological samples compared with many competing biophysical techniques. Improvements in detector and dye technologies coupled to advances in image analysis methods have fuelled recent development towards single-molecule fluorescence microscopy, which can utilize light microscopy tools to enable the faithful detection and analysis of single fluorescent molecules used as reporter tags in biological samples. For example, the discovery of GFP, initiating the so-called ‘green revolution’, has pushed experimental tools in the biosciences to a completely new level of functional imaging of living samples, culminating in single fluorescent protein molecule detection. Today, fluorescence microscopy is an indispensable tool in single-molecule investigations, providing a high signal-to-noise ratio for visualization while still retaining the key features in the physiological context of native biological systems. In this review, we discuss some of the recent discoveries in the life sciences which have been enabled using single-molecule fluorescence microscopy, paying particular attention to the so-called ‘super-resolution’ fluorescence microscopy techniques in live cells, which are at the cutting-edge of these methods. In particular, how these tools can reveal new insights into long-standing puzzles in biology: old problems, which have been impossible to tackle using other more traditional tools until the emergence of new single-molecule fluorescence microscopy techniques. PMID:28694303

Dictionary of Microscopy

NASA Astrophysics Data System (ADS)

Heath, Julian

2005-10-01

The past decade has seen huge advances in the application of microscopy in all areas of science. This welcome development in microscopy has been paralleled by an expansion of the vocabulary of technical terms used in microscopy: terms have been coined for new instruments and techniques and, as microscopes reach even higher resolution, the use of terms that relate to the optical and physical principles underpinning microscopy is now commonplace. The Dictionary of Microscopy was compiled to meet this challenge and provides concise definitions of over 2,500 terms used in the fields of light microscopy, electron microscopy, scanning probe microscopy, x-ray microscopy and related techniques. Written by Dr Julian P. Heath, Editor of Microscopy and Analysis, the dictionary is intended to provide easy navigation through the microscopy terminology and to be a first point of reference for definitions of new and established terms. The Dictionary of Microscopy is an essential, accessible resource for: students who are new to the field and are learning about microscopes equipment purchasers who want an explanation of the terms used in manufacturers' literature scientists who are considering using a new microscopical technique experienced microscopists as an aide mémoire or quick source of reference librarians, the press and marketing personnel who require definitions for technical reports.

A compilation of cold cases using scanning electron microscopy at the University of Rhode Island

NASA Astrophysics Data System (ADS)

Platek, Michael J.; Gregory, Otto J.

2015-10-01

Scanning electron microscopy combined with microchemical analysis has evolved into one of the most widely used instruments in forensic science today. In particular, the environmental scanning electron microscope (SEM) in conjunction with energy dispersive spectroscopy (EDS), has created unique opportunities in forensic science in regard to the examination of trace evidence; i.e. the examination of evidence without altering the evidence with conductive coatings, thereby enabling criminalists to solve cases that were previously considered unsolvable. Two cold cases were solved at URI using a JEOL 5900 LV SEM in conjunction with EDS. A cold case murder and a cold missing person case will be presented from the viewpoint of the microscopist and will include sample preparation, as well as image and chemical analysis of the trace evidence using electron microscopy and optical microscopy.

[Watching dance of the molecules - CARS microscopy].

PubMed

Korczyński, Jaroslaw; Kubiak, Katarzyna; Węgłowska, Edyta

2017-01-01

CARS (Coherent Anti-Stokes Raman Scattering) microscopy is an imaging method for living cells visualization as well as for food or cosmetics material analysis without the need for staining. The near infrared laser source generates the CARS signal - the characteristic intrinsic vibrational contrast of the molecules in a sample which is no longer caused by staining, but by the molecules themselves. It provides the benefit of a non-toxic, non-destructive and almost noninvasive method for sample imaging. CARS can easily be combined with fluorescence confocal microscopy so it is an excellent complementary imaging method. In this article we showed some of the applications for this technology: imaging of lipid droplets inside human HaCaT cells and analysis of the composition of cosmetic products. Moreover we believe, that soon new fields of application become accessible for this rapidly developing branch of microscopy.

(Machine-)Learning to analyze in vivo microscopy: Support vector machines.

PubMed

Wang, Michael F Z; Fernandez-Gonzalez, Rodrigo

2017-11-01

The development of new microscopy techniques for super-resolved, long-term monitoring of cellular and subcellular dynamics in living organisms is revealing new fundamental aspects of tissue development and repair. However, new microscopy approaches present several challenges. In addition to unprecedented requirements for data storage, the analysis of high resolution, time-lapse images is too complex to be done manually. Machine learning techniques are ideally suited for the (semi-)automated analysis of multidimensional image data. In particular, support vector machines (SVMs), have emerged as an efficient method to analyze microscopy images obtained from animals. Here, we discuss the use of SVMs to analyze in vivo microscopy data. We introduce the mathematical framework behind SVMs, and we describe the metrics used by SVMs and other machine learning approaches to classify image data. We discuss the influence of different SVM parameters in the context of an algorithm for cell segmentation and tracking. Finally, we describe how the application of SVMs has been critical to study protein localization in yeast screens, for lineage tracing in C. elegans, or to determine the developmental stage of Drosophila embryos to investigate gene expression dynamics. We propose that SVMs will become central tools in the analysis of the complex image data that novel microscopy modalities have made possible. This article is part of a Special Issue entitled: Biophysics in Canada, edited by Lewis Kay, John Baenziger, Albert Berghuis and Peter Tieleman. Copyright © 2017 Elsevier B.V. All rights reserved.

Not all that glitters is gold-Electron microscopy study on uptake of gold nanoparticles in Daphnia magna and related artifacts.

PubMed

Jensen, Louise Helene Søgaard; Skjolding, Lars Michael; Thit, Amalie; Sørensen, Sara Nørgaard; Købler, Carsten; Mølhave, Kristian; Baun, Anders

2017-06-01

Increasing use of engineered nanoparticles has led to extensive research into their potential hazards to the environment and human health. Cellular uptake from the gut is sparsely investigated, and microscopy techniques applied for uptake studies can result in misinterpretations. Various microscopy techniques were used to investigate internalization of 10-nm gold nanoparticles in Daphnia magna gut lumen and gut epithelial cells following 24-h exposure and outline potential artifacts (i.e., high-contrast precipitates from sample preparation related to these techniques). Light sheet microscopy confirmed accumulation of gold nanoparticles in the gut lumen. Scanning transmission electron microscopy and elemental analysis revealed gold nanoparticles attached to the microvilli of gut cells. Interestingly, the peritrophic membrane appeared to act as a semipermeable barrier between the lumen and the gut epithelium, permitting only single particles through. Structures resembling nanoparticles were also observed inside gut cells. Elemental analysis could not verify these to be gold, and they were likely artifacts from the preparation, such as osmium and iron. Importantly, gold nanoparticles were found inside holocrine cells with disrupted membranes. Thus, false-positive observations of nanoparticle internalization may result from either preparation artifacts or mistaking disrupted cells for intact cells. These findings emphasize the importance of cell integrity and combining elemental analysis with the localization of internalized nanoparticles using transmission electron microscopy. Environ Toxicol Chem 2017;36:1503-1509. © 2016 SETAC. © 2016 SETAC.

Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

NASA Astrophysics Data System (ADS)

Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

2010-05-01

Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

Single cell elemental analysis using nuclear microscopy

NASA Astrophysics Data System (ADS)

Ren, M. Q.; Thong, P. S. P.; Kara, U.; Watt, F.

1999-04-01

The use of Particle Induced X-ray Emission (PIXE), Rutherford Backscattering Spectrometry (RBS) and Scanning Transmission Ion Microscopy (STIM) to provide quantitative elemental analysis of single cells is an area which has high potential, particularly when the trace elements such as Ca, Fe, Zn and Cu can be monitored. We describe the methodology of sample preparation for two cell types, the procedures of cell imaging using STIM, and the quantitative elemental analysis of single cells using RBS and PIXE. Recent work on single cells at the Nuclear Microscopy Research Centre,National University of Singapore has centred around two research areas: (a) Apoptosis (programmed cell death), which has been recently implicated in a wide range of pathological conditions such as cancer, Parkinson's disease etc, and (b) Malaria (infection of red blood cells by the malaria parasite). Firstly we present results on the elemental analysis of human Chang liver cells (ATTCC CCL 13) where vanadium ions were used to trigger apoptosis, and demonstrate that nuclear microscopy has the capability of monitoring vanadium loading within individual cells. Secondly we present the results of elemental changes taking place in individual mouse red blood cells which have been infected with the malaria parasite and treated with the anti-malaria drug Qinghaosu (QHS).

Characterization of Homopolymer and Polymer Blend Films by Phase Sensitive Acoustic Microscopy

NASA Astrophysics Data System (ADS)

Ngwa, Wilfred; Wannemacher, Reinhold; Grill, Wolfgang

2003-03-01

CHARACTERIZATION OF HOMOPOLYMER AND POLYMER BLEND FILMS BY PHASE SENSITIVE ACOUSTIC MICROSCOPY W Ngwa, R Wannemacher, W Grill Institute of Experimental Physics II, University of Leipzig, 04103 Leipzig, Germany Abstract We have used phase sensitive acoustic microscopy (PSAM) to study homopolymer thin films of polystyrene (PS) and poly (methyl methacrylate) (PMMA), as well as PS/PMMA blend films. We show from our results that PSAM can be used as a complementary and highly valuable technique for elucidating the three-dimensional (3D) morphology and micromechanical properties of thin films. Three-dimensional image acquisition with vector contrast provides the basis for: complex V(z) analysis (per image pixel), 3D image processing, height profiling, and subsurface image analysis of the polymer films. Results show good agreement with previous studies. In addition, important new information on the three dimensional structure and properties of polymer films is obtained. Homopolymer film structure analysis reveals (pseudo-) dewetting by retraction of droplets, resulting in a morphology that can serve as a starting point for the analysis of polymer blend thin films. The outcome of confocal laser scanning microscopy studies, performed on the same samples are correlated with the obtained results. Advantages and limitations of PSAM are discussed.

Evaluation of agave fiber delignification by means of microscopy techniques and image analysis.

PubMed

Hernández-Hernández, Hilda M; Chanona-Pérez, Jorge J; Calderón-Domínguez, Georgina; Perea-Flores, María J; Mendoza-Pérez, Jorge A; Vega, Alberto; Ligero, Pablo; Palacios-González, Eduardo; Farrera-Rebollo, Reynold R

2014-10-01

Recently, the use of different types of natural fibers to produce paper and textiles from agave plants has been proposed. Agave atrovirens can be a good source of cellulose and lignin; nevertheless, the microstructural changes that happen during delignification have scarcely been studied. The aim of this work was to study the microstructural changes that occur during the delignification of agave fibers by means of microscopy techniques and image analysis. The fibers of A. atrovirens were obtained from leaves using convective drying, milling, and sieving. Fibers were processed using the Acetosolv pulping method at different concentrations of acetic acid; increasing acid concentration promoted higher levels of delignification, structural damage, and the breakdown of fiber clumps. Delignification followed by spectrometric analysis and microstructural studies were carried out by light, confocal laser scanning and scanning electron microscopy and showed that the delignification process follows three stages: initial, bulk, and residual. Microscopy techniques and image analysis were efficient tools for microstructural characterization during delignification of agave fibers, allowing quantitative evaluation of the process and the development of linear prediction models. The data obtained integrated numerical and microstructural information that could be valuable for the study of pulping of lignocellulosic materials.

Histological Analysis of the Arabidopsis Gynoecium and Ovules Using Chloral Hydrate Clearing and Differential Interference Contrast Light Microscopy.

PubMed

Franks, Robert G

2016-01-01

The use of chloral hydrate optical clearing paired with differential interference contrast microscopy allows the analysis of internal structures of developing plant organs without the need for paraffin embedding and sectioning. This approach is appropriate for the analysis of the developing gynoecium or seedpod of the flowering plant Arabidopsis thaliana and many other types of fixed plant material. Early stages of ovule development are observable with this approach.

Qualitative and quantitative interpretation of SEM image using digital image processing.

PubMed

Saladra, Dawid; Kopernik, Magdalena

2016-10-01

The aim of the this study is improvement of qualitative and quantitative analysis of scanning electron microscope micrographs by development of computer program, which enables automatic crack analysis of scanning electron microscopy (SEM) micrographs. Micromechanical tests of pneumatic ventricular assist devices result in a large number of micrographs. Therefore, the analysis must be automatic. Tests for athrombogenic titanium nitride/gold coatings deposited on polymeric substrates (Bionate II) are performed. These tests include microshear, microtension and fatigue analysis. Anisotropic surface defects observed in the SEM micrographs require support for qualitative and quantitative interpretation. Improvement of qualitative analysis of scanning electron microscope images was achieved by a set of computational tools that includes binarization, simplified expanding, expanding, simple image statistic thresholding, the filters Laplacian 1, and Laplacian 2, Otsu and reverse binarization. Several modifications of the known image processing techniques and combinations of the selected image processing techniques were applied. The introduced quantitative analysis of digital scanning electron microscope images enables computation of stereological parameters such as area, crack angle, crack length, and total crack length per unit area. This study also compares the functionality of the developed computer program of digital image processing with existing applications. The described pre- and postprocessing may be helpful in scanning electron microscopy and transmission electron microscopy surface investigations. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

Analysis of Particulate and Fiber Debris Samples Returned from the International Space Station

NASA Technical Reports Server (NTRS)

Perry, Jay L.; Coston, James E.

2014-01-01

During the period of International Space Station (ISS) Increments 30 and 31, crewmember reports cited differences in the cabin environment relating to particulate matter and fiber debris compared to earlier experience as well as allergic responses to the cabin environment. It was hypothesized that a change in the cabin atmosphere's suspended particulate matter load may be responsible for the reported situation. Samples were collected and returned to ground-based laboratories for assessment. Assessments included physical classification, optical microscopy and photographic analysis, and scanning electron microscopy (SEM) evaluation using energy dispersive X-ray spectrometry (EDS) methods. Particular points of interest for assessing the samples were for the presence of allergens, carbon dioxide removal assembly (CDRA) zeolite dust, and FGB panel fibers. The results from the physical classification, optical microscopy and photographic analysis, and SEM EDS analysis are presented and discussed.

Qualitative and quantitative analysis of PMN/T-cell interactions by InFlow and super-resolution microscopy.

PubMed

Balta, Emre; Stopp, Julian; Castelletti, Laura; Kirchgessner, Henning; Samstag, Yvonne; Wabnitz, Guido H

2017-01-01

Neutrophils or polymorphonuclear cells (PMN) eliminate bacteria via phagocytosis and/or NETosis. Apart from these conventional roles, PMN also have immune-regulatory functions. They can transdifferentiate and upregulate MHCII as well as ligands for costimulatory receptors which enables them to behave as antigen presenting cells (APC). The initial step for activating T-cells is the formation of an immune synapse between T-cells and antigen-presenting cells. However, the immune synapse that develops at the PMN/T-cell contact zone is as yet hardly investigated due to the non-availability of methods for analysis of large number of PMN interactions. In order to overcome these obstacles, we introduce here a workflow to analyse the immune synapse of primary human PMN and T-cells using multispectral imaging flow cytometry (InFlow microscopy) and super-resolution microscopy. For that purpose, we used CD3 and CD66b as the lineage markers for T-cells and PMN, respectively. Thereafter, we applied and critically discussed various "masks" for identification of T-cell PMN interactions. Using this approach, we found that a small fraction of transdifferentiated PMN (CD66b + CD86 high ) formed stable PMN/T-cell conjugates. Interestingly, while both CD3 and CD66b accumulation in the immune synapse was dependent on the maturation state of the PMN, only CD3 accumulation was greatly enhanced by the presence of superantigen. The actin cytoskeleton was weakly rearranged at the PMN side on the immune synapse upon contact with a T-cell in the presence of superantigen. A more detailed analysis using super-resolution microscopy (structured-illumination microscopy, SIM) confirmed this finding. Together, we present an InFlow microscopy based approach for the large scale analysis of PMN/T-cell interactions and - combined with SIM - a possibility for an in-depth analysis of protein translocation at the site of interactions. Copyright © 2016 Elsevier Inc. All rights reserved.

Segmentation and Morphometric Analysis of Cells from Fluorescence Microscopy Images of Cytoskeletons

PubMed Central

Ujihara, Yoshihiro; Nakamura, Masanori; Miyazaki, Hiroshi; Wada, Shigeo

2013-01-01

We developed a method to reconstruct cell geometry from confocal fluorescence microscopy images of the cytoskeleton. In the method, region growing was implemented twice. First, it was applied to the extracellular regions to differentiate them from intracellular noncytoskeletal regions, which both appear black in fluorescence microscopy imagery, and then to cell regions for cell identification. Analysis of morphological parameters revealed significant changes in cell shape associated with cytoskeleton disruption, which offered insight into the mechanical role of the cytoskeleton in maintaining cell shape. The proposed segmentation method is promising for investigations on cell morphological changes with respect to internal cytoskeletal structures. PMID:23762186

Characterization of inertial confinement fusion (ICF) targets using PIXE, RBS, and STIM analysis.

PubMed

Li, Yongqiang; Liu, Xue; Li, Xinyi; Liu, Yiyang; Zheng, Yi; Wang, Min; Shen, Hao

2013-08-01

Quality control of the inertial confinement fusion (ICF) target in the laser fusion program is vital to ensure that energy deposition from the lasers results in uniform compression and minimization of Rayleigh-Taylor instabilities. The technique of nuclear microscopy with ion beam analysis is a powerful method to provide characterization of ICF targets. Distribution of elements, depth profile, and density image of ICF targets can be identified by particle-induced X-ray emission, Rutherford backscattering spectrometry, and scanning transmission ion microscopy. We present examples of ICF target characterization by nuclear microscopy at Fudan University in order to demonstrate their potential impact in assessing target fabrication processes.

Application of He ion microscopy for material analysis

NASA Astrophysics Data System (ADS)

Altmann, F.; Simon, M.; Klengel, R.

2009-05-01

Helium ion beam microscopy (HIM) is a new high resolution imaging technique. The use of Helium ions instead of electrons enables none destructive imaging combined with contrasts quite similar to that from Gallium ion beam imaging. The use of very low probe currents and the comfortable charge compensation using low energy electrons offer imaging of none conductive samples without conductive coating. An ongoing microelectronic sample with Gold/Aluminum interconnects and polymer electronic devices were chosen to evaluate HIM in comparison to scanning electron microscopy (SEM). The aim was to look for key applications of HIM in material analysis. Main focus was on complementary contrast mechanisms and imaging of none conductive samples.

Segmentation and morphometric analysis of cells from fluorescence microscopy images of cytoskeletons.

PubMed

Ujihara, Yoshihiro; Nakamura, Masanori; Miyazaki, Hiroshi; Wada, Shigeo

2013-01-01

We developed a method to reconstruct cell geometry from confocal fluorescence microscopy images of the cytoskeleton. In the method, region growing was implemented twice. First, it was applied to the extracellular regions to differentiate them from intracellular noncytoskeletal regions, which both appear black in fluorescence microscopy imagery, and then to cell regions for cell identification. Analysis of morphological parameters revealed significant changes in cell shape associated with cytoskeleton disruption, which offered insight into the mechanical role of the cytoskeleton in maintaining cell shape. The proposed segmentation method is promising for investigations on cell morphological changes with respect to internal cytoskeletal structures.

Quantitative super-resolution single molecule microscopy dataset of YFP-tagged growth factor receptors.

PubMed

Lukeš, Tomáš; Pospíšil, Jakub; Fliegel, Karel; Lasser, Theo; Hagen, Guy M

2018-03-01

Super-resolution single molecule localization microscopy (SMLM) is a method for achieving resolution beyond the classical limit in optical microscopes (approx. 200 nm laterally). Yellow fluorescent protein (YFP) has been used for super-resolution single molecule localization microscopy, but less frequently than other fluorescent probes. Working with YFP in SMLM is a challenge because a lower number of photons are emitted per molecule compared with organic dyes, which are more commonly used. Publically available experimental data can facilitate development of new data analysis algorithms. Four complete, freely available single molecule super-resolution microscopy datasets on YFP-tagged growth factor receptors expressed in a human cell line are presented, including both raw and analyzed data. We report methods for sample preparation, for data acquisition, and for data analysis, as well as examples of the acquired images. We also analyzed the SMLM datasets using a different method: super-resolution optical fluctuation imaging (SOFI). The 2 modes of analysis offer complementary information about the sample. A fifth single molecule super-resolution microscopy dataset acquired with the dye Alexa 532 is included for comparison purposes. This dataset has potential for extensive reuse. Complete raw data from SMLM experiments have typically not been published. The YFP data exhibit low signal-to-noise ratios, making data analysis a challenge. These datasets will be useful to investigators developing their own algorithms for SMLM, SOFI, and related methods. The data will also be useful for researchers investigating growth factor receptors such as ErbB3.

Revista de Documentacao de Estudos em Linguistica Teorica e Aplicada (DELTA): Novos Estudos em Gamatica Gerativa (Journal of Documentary Studies in Theoretical and Applied Linguistics [DELTA]: New Studies in Generative Grammar).

ERIC Educational Resources Information Center

Revista de Documentacao de Estudos em Linguistica Teorica e Aplicada, 2000

2000-01-01

This issue contains the following articles: "Resumption and Last Resort" (Joseph Aoun); "Existentials, A-Chains, and Reconstruction" (Norbert Hornstein); "How Long Was the Nineteenth Century" (David Lightfoot); "Formal Features and Parameter Setting: A View From Portuguese Past Participles and Romance Future…

Maximizing the Biochemical Resolving Power of Fluorescence Microscopy

PubMed Central

Esposito, Alessandro; Popleteeva, Marina; Venkitaraman, Ashok R.

2013-01-01

Most recent advances in fluorescence microscopy have focused on achieving spatial resolutions below the diffraction limit. However, the inherent capability of fluorescence microscopy to non-invasively resolve different biochemical or physical environments in biological samples has not yet been formally described, because an adequate and general theoretical framework is lacking. Here, we develop a mathematical characterization of the biochemical resolution in fluorescence detection with Fisher information analysis. To improve the precision and the resolution of quantitative imaging methods, we demonstrate strategies for the optimization of fluorescence lifetime, fluorescence anisotropy and hyperspectral detection, as well as different multi-dimensional techniques. We describe optimized imaging protocols, provide optimization algorithms and describe precision and resolving power in biochemical imaging thanks to the analysis of the general properties of Fisher information in fluorescence detection. These strategies enable the optimal use of the information content available within the limited photon-budget typically available in fluorescence microscopy. This theoretical foundation leads to a generalized strategy for the optimization of multi-dimensional optical detection, and demonstrates how the parallel detection of all properties of fluorescence can maximize the biochemical resolving power of fluorescence microscopy, an approach we term Hyper Dimensional Imaging Microscopy (HDIM). Our work provides a theoretical framework for the description of the biochemical resolution in fluorescence microscopy, irrespective of spatial resolution, and for the development of a new class of microscopes that exploit multi-parametric detection systems. PMID:24204821

Cell segmentation in phase contrast microscopy images via semi-supervised classification over optics-related features.

PubMed

Su, Hang; Yin, Zhaozheng; Huh, Seungil; Kanade, Takeo

2013-10-01

Phase-contrast microscopy is one of the most common and convenient imaging modalities to observe long-term multi-cellular processes, which generates images by the interference of lights passing through transparent specimens and background medium with different retarded phases. Despite many years of study, computer-aided phase contrast microscopy analysis on cell behavior is challenged by image qualities and artifacts caused by phase contrast optics. Addressing the unsolved challenges, the authors propose (1) a phase contrast microscopy image restoration method that produces phase retardation features, which are intrinsic features of phase contrast microscopy, and (2) a semi-supervised learning based algorithm for cell segmentation, which is a fundamental task for various cell behavior analysis. Specifically, the image formation process of phase contrast microscopy images is first computationally modeled with a dictionary of diffraction patterns; as a result, each pixel of a phase contrast microscopy image is represented by a linear combination of the bases, which we call phase retardation features. Images are then partitioned into phase-homogeneous atoms by clustering neighboring pixels with similar phase retardation features. Consequently, cell segmentation is performed via a semi-supervised classification technique over the phase-homogeneous atoms. Experiments demonstrate that the proposed approach produces quality segmentation of individual cells and outperforms previous approaches. Copyright © 2013 Elsevier B.V. All rights reserved.

Analysis of Long Bone and Vertebral Failure Patterns.

DTIC Science & Technology

1982-09-30

processes further supported the findings of • :the scanning electron microscopy studies . In the impacted animals, the cartilage surface was eroded... cartilage matrix. In the six years post-impaction group, the articular cartilage had converted to fibrocartilage instead of normal hyaline cartilage . The...columns of four rhesus monkeys have been collected and are being processed for study with light microscopy and scanning electron microscopy. The baboon

A Quantitative Approach to Scar Analysis

PubMed Central

Khorasani, Hooman; Zheng, Zhong; Nguyen, Calvin; Zara, Janette; Zhang, Xinli; Wang, Joyce; Ting, Kang; Soo, Chia

2011-01-01

Analysis of collagen architecture is essential to wound healing research. However, to date no consistent methodologies exist for quantitatively assessing dermal collagen architecture in scars. In this study, we developed a standardized approach for quantitative analysis of scar collagen morphology by confocal microscopy using fractal dimension and lacunarity analysis. Full-thickness wounds were created on adult mice, closed by primary intention, and harvested at 14 days after wounding for morphometrics and standard Fourier transform-based scar analysis as well as fractal dimension and lacunarity analysis. In addition, transmission electron microscopy was used to evaluate collagen ultrastructure. We demonstrated that fractal dimension and lacunarity analysis were superior to Fourier transform analysis in discriminating scar versus unwounded tissue in a wild-type mouse model. To fully test the robustness of this scar analysis approach, a fibromodulin-null mouse model that heals with increased scar was also used. Fractal dimension and lacunarity analysis effectively discriminated unwounded fibromodulin-null versus wild-type skin as well as healing fibromodulin-null versus wild-type wounds, whereas Fourier transform analysis failed to do so. Furthermore, fractal dimension and lacunarity data also correlated well with transmission electron microscopy collagen ultrastructure analysis, adding to their validity. These results demonstrate that fractal dimension and lacunarity are more sensitive than Fourier transform analysis for quantification of scar morphology. PMID:21281794

Cost effectiveness of OptiMal® rapid diagnostic test for malaria in remote areas of the Amazon Region, Brazil

PubMed Central

2010-01-01

Background In areas with limited structure in place for microscopy diagnosis, rapid diagnostic tests (RDT) have been demonstrated to be effective. Method The cost-effectiveness of the Optimal® and thick smear microscopy was estimated and compared. Data were collected on remote areas of 12 municipalities in the Brazilian Amazon. Data sources included the National Malaria Control Programme of the Ministry of Health, the National Healthcare System reimbursement table, hospitalization records, primary data collected from the municipalities, and scientific literature. The perspective was that of the Brazilian public health system, the analytical horizon was from the start of fever until the diagnostic results provided to patient and the temporal reference was that of year 2006. The results were expressed in costs per adequately diagnosed cases in 2006 U.S. dollars. Sensitivity analysis was performed considering key model parameters. Results In the case base scenario, considering 92% and 95% sensitivity for thick smear microscopy to Plasmodium falciparum and Plasmodium vivax, respectively, and 100% specificity for both species, thick smear microscopy is more costly and more effective, with an incremental cost estimated at US$549.9 per adequately diagnosed case. In sensitivity analysis, when sensitivity and specificity of microscopy for P. vivax were 0.90 and 0.98, respectively, and when its sensitivity for P. falciparum was 0.83, the RDT was more cost-effective than microscopy. Conclusion Microscopy is more cost-effective than OptiMal® in these remote areas if high accuracy of microscopy is maintained in the field. Decision regarding use of rapid tests for diagnosis of malaria in these areas depends on current microscopy accuracy in the field. PMID:20937094

Comparison of loop-mediated isothermal amplification assay and smear microscopy with culture for the diagnostic accuracy of tuberculosis.

PubMed

Gelaw, Baye; Shiferaw, Yitayal; Alemayehu, Marta; Bashaw, Abate Assefa

2017-01-17

Tuberculosis (TB) caused by Mycobacterium tuberculosis is one of the leading causes of death from infectious diseases worldwide. Sputum smear microscopy remains the most widely available pulmonary TB diagnostic tool particularly in resource limited settings. A highly sensitive diagnostic with minimal infrastructure, cost and training is required. Hence, we assessed the diagnostic performance of Loop-mediated isothermal amplification (LAMP) assay in detecting M.tuberculosis infection in sputum sample compared to LED fluorescent smear microscopy and culture. A cross-sectional study was conducted at the University of Gondar Hospital from June 01, 2015 to August 30, 2015. Pulmonary TB diagnosis using sputum LED fluorescence smear microscopy, TB-LAMP assay and culture were done. A descriptive analysis was used to determine demographic characteristics of the study participants. Analysis of sensitivity and specificity for smear microscopy and TB-LAMP compared with culture as a reference test was performed. Cohen's kappa was calculated as a measure of agreement between the tests. A total of 78 pulmonary presumptive TB patients sputum sample were analyzed. The overall sensitivity and specificity of LAMP were 75 and 98%, respectively. Among smear negative sputum samples, 33.3% sensitivity and 100% specificity of LAMP were observed. Smear microscopy showed 78.6% sensitivity and 98% specificity. LAMP and smear in series had sensitivity of 67.8% and specificity of 100%. LAMP and smear in parallel had sensitivity of 85.7% and specificity of 96%. The agreement between LAMP and fluorescent smear microscopy tests was very good (κ = 0.83, P-value ≤0.0001). TB-LAMP showed similar specificity but a slightly lower sensitivity with LED fluorescence microscopy. The specificity of LAMP and smear microscopy in series was high. The sensitivity of LAMP was insufficient for smear negative sputum samples.

Microbiologically influenced corrosion: looking to the future.

PubMed

Videla, Héctor A; Herrera, Liz K

2005-09-01

This review discusses the state-of-the-art of research into biocorrosion and the biofouling of metals and alloys of industrial usage. The key concepts needed to understand the main effects of microorganisms on metal decay, and current trends in monitoring and control strategies to mitigate the deleterious effects of biocorrosion and biofouling are also described. Several relevant cases of biocorrosion studied by our research group are provided as examples: (i) biocorrosion of aluminum and its alloys by fungal contaminants of jet fuels; (ii) sulfate-reducing bacteria (SRB)-induced corrosion of steel; (iii) biocorrosion and biofouling interactions in the marine environment; (iv) monitoring strategies for assessing biocorrosion in industrial water systems; (v) microbial inhibition of corrosion; (vi) use and limitations of electrochemical techniques for evaluating biocorrosion effects. Future prospects in the field are described with respect to the potential of innovative techniques in microscopy (environmental scanning electron microscopy, confocal scanning laser microscopy, atomic force microscopy), new spectroscopic techniques for the study of corrosion products and biofilms (energy dispersion X-ray analysis, X-ray photoelectron spectroscopy, electron microprobe analysis) and electrochemistry (electrochemical impedance spectroscopy, electrochemical noise analysis).

Determination of precursor sites for pitting corrosion of polycrystalline titanium by using different techniques

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garfias-Mesias, L.F.; Alodan, M.; James, P.I.

1998-06-01

Scanning electrochemical microscopy (SECM) in ferrocyanide and bromide solutions was used to locate active sites (pitting precursors) on polycrystalline Ti where oxidation of Br{sup {minus}} and Fe(CN){sub 6}{sup 4{minus}} was possible. Analysis of the electrochemically active sites was done by using electron microscopy (SEM), energy dispersive X-ray analysis (EDX), atomic force microscopy (AFM), and in situ confocal laser scanning microscopy (CLSM). In most cases, the active sites were found to be associated with particles (inclusions) which contained mainly Al and Si; however, some other areas not associated with particles were also found to be active. Although the size of themore » inclusions was normally smaller than 20 {micro}m, as revealed by SEM and AFM imaging, in some cases larger particles were also found. Pitting corrosion tests in bromide solution at potentials above 1.5 V{sub SCE} followed by EDX analysis inside the pits and in situ CLSM observation, confirmed that most of the localized attack started in the areas where particles had been located.« less

Development of a viability standard curve for microencapsulated probiotic bacteria using confocal microscopy and image analysis software.

PubMed

Moore, Sarah; Kailasapathy, Kasipathy; Phillips, Michael; Jones, Mark R

2015-07-01

Microencapsulation is proposed to protect probiotic strains from food processing procedures and to maintain probiotic viability. Little research has described the in situ viability of microencapsulated probiotics. This study successfully developed a real-time viability standard curve for microencapsulated bacteria using confocal microscopy, fluorescent dyes and image analysis software. Copyright © 2015 Elsevier B.V. All rights reserved.

The effects of viscoelastic polymer substrates on adult stem cell differentiation

NASA Astrophysics Data System (ADS)

Chang, Chungchueh; Fields, Adam; Ramek, Alex; Jurukovski, Vladimir; Simon, Marcia; Rafailovich, Miriam

2009-03-01

Dental Pulp Stem Cells (DPSCs) are known to differentiate in either bone, dentine, or nerve tissue by different environment signals. In this study, we have determined whether differentiation could only through modification of the substrate mechanics. Atomic Force Microscopy (AFM) on Shear Modulation Force Microscopy (SMFM) mode indicated that the spun-cast polybutadiene (PB) thin films could be used to provide different stiffness substrates by changing the thicknesses of thin films. DPSCs were then plated on these substrates and cultured in standard media. After 28 days incubation, Lasar Scanning Confocal Microscopy (LSCM) with mercury lamp indicated that the crystals were observed only on hard surfaces. The Scanning Electron Microscopy (SEM) and Energy Dispersive X-ray analysis (EDX analysis) indicated that the crystals are calcium phosphates. The Glancing Incidence Diffraction (GID) was also used to determine the structure of crystals. These results indicate that DPSCs could be differentiated into osteoblasts by mechanical stimuli from substrate mechanics.

Gaps analysis for CD metrology beyond the 22nm node

NASA Astrophysics Data System (ADS)

Bunday, Benjamin; Germer, Thomas A.; Vartanian, Victor; Cordes, Aaron; Cepler, Aron; Settens, Charles

2013-04-01

This paper will examine the future for critical dimension (CD) metrology. First, we will present the extensive list of applications for which CD metrology solutions are needed, showing commonalities and differences among the various applications. We will then report on the expected technical limits of the metrology solutions currently being investigated by SEMATECH and others in the industry to address the metrology challenges of future nodes, including conventional CD scanning electron microscopy (CD-SEM) and optical critical dimension (OCD) metrology and new potential solutions such as He-ion microscopy (HeIM, sometimes elsewhere referred to as HIM), CD atomic force microscopy (CD-AFM), CD small-angle x-ray scattering (CD-SAXS), high-voltage scanning electron microscopy (HV-SEM), and other types. A technical gap analysis matrix will then be demonstrated, showing the current state of understanding of the future of the CD metrology space.

Peculiarities of studying an isolated neuron by the method of laser interference microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yusipovich, Alexander I; Kazakova, Tatiana A; Erokhova, Liudmila A

2006-09-30

Actual aspects of using a new method of laser interference microscopy (LIM) for studying nerve cells are discussed. The peculiarities of the LIM display of neurons are demonstrated by the example of isolated neurons of a pond snail Lymnaea stagnalis. A comparative analysis of the images of the cell and subcellular structures of a neuron obtained by the methods of interference microscopy, optical transmission microscopy, and confocal microscopy is performed. Various aspects of the application of LIM for studying the lateral dimensions and internal structure of the cytoplasm and organelles of a neuron in cytology and cell physiology are discussed.more » (laser biology)« less

In Situ and In Vivo Molecular Analysis by Coherent Raman Scattering Microscopy

PubMed Central

Liao, Chien-Sheng; Cheng, Ji-Xin

2017-01-01

Coherent Raman scattering (CRS) microscopy is a high-speed vibrational imaging platform with the ability to visualize the chemical content of a living specimen by using molecular vibrational fingerprints. We review technical advances and biological applications of CRS microscopy. The basic theory of CRS and the state-of-the-art instrumentation of a CRS microscope are presented. We further summarize and compare the algorithms that are used to separate the Raman signal from the nonresonant background, to denoise a CRS image, and to decompose a hyperspectral CRS image into concentration maps of principal components. Important applications of single-frequency and hyperspectral CRS microscopy are highlighted. Potential directions of CRS microscopy are discussed. PMID:27306307

A simple 2D composite image analysis technique for the crystal growth study of L-ascorbic acid.

PubMed

Kumar, Krishan; Kumar, Virender; Lal, Jatin; Kaur, Harmeet; Singh, Jasbir

2017-06-01

This work was destined for 2D crystal growth studies of L-ascorbic acid using the composite image analysis technique. Growth experiments on the L-ascorbic acid crystals were carried out by standard (optical) microscopy, laser diffraction analysis, and composite image analysis. For image analysis, the growth of L-ascorbic acid crystals was captured as digital 2D RGB images, which were then processed to composite images. After processing, the crystal boundaries emerged as white lines against the black (cancelled) background. The crystal boundaries were well differentiated by peaks in the intensity graphs generated for the composite images. The lengths of crystal boundaries measured from the intensity graphs of composite images were in good agreement (correlation coefficient "r" = 0.99) with the lengths measured by standard microscopy. On the contrary, the lengths measured by laser diffraction were poorly correlated with both techniques. Therefore, the composite image analysis can replace the standard microscopy technique for the crystal growth studies of L-ascorbic acid. © 2017 Wiley Periodicals, Inc.

Revista de Documentacao de Estudos em Linguistica Teorica e Aplicada (DELTA): Novos Estudos em Gamatica Gerativa, 2001 (Journal of Documentary Studies in Theoretical and Applied Linguistics [DELTA]: New Studies in Generative Grammar, 2001).

ERIC Educational Resources Information Center

Barbara, Leila, Ed.; Rajagopalan, Kanavillil, Ed.

2001-01-01

These two issues of volume 17, include the following articles: "The Competing Motivation Model in the Functional Domains of Negation" (M. Angelica Furtado da Cunha); "Discursive Resonance and Politeness in Reading and Writing Practices" (Silvana Serrani Infante); "The Acquisition of Relative Clauses in Brazilian…

O que bilíngues bimodais têm a nos dizer sobre desenvolvimento bilíngue?

PubMed Central

de Quadros, Ronice Müller; Lillo-Martin, Diane; Pichler, Deborah Chen

2013-01-01

O objetivo deste trabalho é apresentar o que as pesquisas que estamos desenvolvendo com crianças ouvintes, filhas de pais surdos, adquirindo Língua Brasileira de Sinais (Libras) e Português e Língua de Sinais Americana (ASL) e Inglês (Lillo-Martin et al. 2010) têm a nos dizer sobre desenvolvimento bilíngue. Os dados deste estudo fazem parte de um banco de dados de interações espontâneas coletadas longitudinalmente, alternando contextos de aquisição da Libras e do português como língua alvo, no Brasil e dados coletados longitudinalmente. nos mesmos contextos, de crianças adquirindo ASL e inglês1. Além disso, há também dados do estudo experimental com testes aplicados nos dois pares de línguas que se agregam ao presente estudo. Uma visão geral dos estudos desenvolvidos sobre a aquisição bilíngue bimodal por crianças ouvintes, filhas de pais surdos, será apresentada e, então, serão expostos alguns aspectos linguísticos deste tipo de aquisição, considerando as discussões sobre aquisição bilíngue a partir da pesquisa realizada. PMID:24431480

Malaria surveillance in the Democratic Republic of the Congo: comparison of microscopy, PCR, and rapid diagnostic test.

PubMed

Doctor, Stephanie M; Liu, Yunhao; Whitesell, Amy; Thwai, Kyaw L; Taylor, Steve M; Janko, Mark; Emch, Michael; Kashamuka, Melchior; Muwonga, Jérémie; Tshefu, Antoinette; Meshnick, Steven R

2016-05-01

Malaria surveillance is critical for control efforts, but diagnostic methods frequently disagree. Here, we compare microscopy, PCR, and a rapid diagnostic test in 7137 samples from children in the Democratic Republic of the Congo using latent class analysis. PCR had the highest sensitivity (94.6%) and microscopy had the lowest (76.7%). Copyright © 2016 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Timmermans, F. J.; Otto, C.

New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemicallymore » or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.« less

A Barcoding Strategy Enabling Higher-Throughput Library Screening by Microscopy.

PubMed

Chen, Robert; Rishi, Harneet S; Potapov, Vladimir; Yamada, Masaki R; Yeh, Vincent J; Chow, Thomas; Cheung, Celia L; Jones, Austin T; Johnson, Terry D; Keating, Amy E; DeLoache, William C; Dueber, John E

2015-11-20

Dramatic progress has been made in the design and build phases of the design-build-test cycle for engineering cells. However, the test phase usually limits throughput, as many outputs of interest are not amenable to rapid analytical measurements. For example, phenotypes such as motility, morphology, and subcellular localization can be readily measured by microscopy, but analysis of these phenotypes is notoriously slow. To increase throughput, we developed microscopy-readable barcodes (MiCodes) composed of fluorescent proteins targeted to discernible organelles. In this system, a unique barcode can be genetically linked to each library member, making possible the parallel analysis of phenotypes of interest via microscopy. As a first demonstration, we MiCoded a set of synthetic coiled-coil leucine zipper proteins to allow an 8 × 8 matrix to be tested for specific interactions in micrographs consisting of mixed populations of cells. A novel microscopy-readable two-hybrid fluorescence localization assay for probing candidate interactions in the cytosol was also developed using a bait protein targeted to the peroxisome and a prey protein tagged with a fluorescent protein. This work introduces a generalizable, scalable platform for making microscopy amenable to higher-throughput library screening experiments, thereby coupling the power of imaging with the utility of combinatorial search paradigms.

Understanding amyloid aggregation by statistical analysis of atomic force microscopy images

NASA Astrophysics Data System (ADS)

Adamcik, Jozef; Jung, Jin-Mi; Flakowski, Jérôme; de Los Rios, Paolo; Dietler, Giovanni; Mezzenga, Raffaele

2010-06-01

The aggregation of proteins is central to many aspects of daily life, including food processing, blood coagulation, eye cataract formation disease and prion-related neurodegenerative infections. However, the physical mechanisms responsible for amyloidosis-the irreversible fibril formation of various proteins that is linked to disorders such as Alzheimer's, Creutzfeldt-Jakob and Huntington's diseases-have not yet been fully elucidated. Here, we show that different stages of amyloid aggregation can be examined by performing a statistical polymer physics analysis of single-molecule atomic force microscopy images of heat-denatured β-lactoglobulin fibrils. The atomic force microscopy analysis, supported by theoretical arguments, reveals that the fibrils have a multistranded helical shape with twisted ribbon-like structures. Our results also indicate a possible general model for amyloid fibril assembly and illustrate the potential of this approach for investigating fibrillar systems.

ThunderSTORM: a comprehensive ImageJ plug-in for PALM and STORM data analysis and super-resolution imaging

PubMed Central

Ovesný, Martin; Křížek, Pavel; Borkovec, Josef; Švindrych, Zdeněk; Hagen, Guy M.

2014-01-01

Summary: ThunderSTORM is an open-source, interactive and modular plug-in for ImageJ designed for automated processing, analysis and visualization of data acquired by single-molecule localization microscopy methods such as photo-activated localization microscopy and stochastic optical reconstruction microscopy. ThunderSTORM offers an extensive collection of processing and post-processing methods so that users can easily adapt the process of analysis to their data. ThunderSTORM also offers a set of tools for creation of simulated data and quantitative performance evaluation of localization algorithms using Monte Carlo simulations. Availability and implementation: ThunderSTORM and the online documentation are both freely accessible at https://code.google.com/p/thunder-storm/ Contact: guy.hagen@lf1.cuni.cz Supplementary information: Supplementary data are available at Bioinformatics online. PMID:24771516

Rapid analysis and exploration of fluorescence microscopy images.

PubMed

Pavie, Benjamin; Rajaram, Satwik; Ouyang, Austin; Altschuler, Jason M; Steininger, Robert J; Wu, Lani F; Altschuler, Steven J

2014-03-19

Despite rapid advances in high-throughput microscopy, quantitative image-based assays still pose significant challenges. While a variety of specialized image analysis tools are available, most traditional image-analysis-based workflows have steep learning curves (for fine tuning of analysis parameters) and result in long turnaround times between imaging and analysis. In particular, cell segmentation, the process of identifying individual cells in an image, is a major bottleneck in this regard. Here we present an alternate, cell-segmentation-free workflow based on PhenoRipper, an open-source software platform designed for the rapid analysis and exploration of microscopy images. The pipeline presented here is optimized for immunofluorescence microscopy images of cell cultures and requires minimal user intervention. Within half an hour, PhenoRipper can analyze data from a typical 96-well experiment and generate image profiles. Users can then visually explore their data, perform quality control on their experiment, ensure response to perturbations and check reproducibility of replicates. This facilitates a rapid feedback cycle between analysis and experiment, which is crucial during assay optimization. This protocol is useful not just as a first pass analysis for quality control, but also may be used as an end-to-end solution, especially for screening. The workflow described here scales to large data sets such as those generated by high-throughput screens, and has been shown to group experimental conditions by phenotype accurately over a wide range of biological systems. The PhenoBrowser interface provides an intuitive framework to explore the phenotypic space and relate image properties to biological annotations. Taken together, the protocol described here will lower the barriers to adopting quantitative analysis of image based screens.

Scanning capacitance microscopy of ErAs nanoparticles embedded in GaAs pn junctions

NASA Astrophysics Data System (ADS)

Park, K. W.; Nair, H. P.; Crook, A. M.; Bank, S. R.; Yu, E. T.

2011-09-01

Scanning capacitance microscopy is used to characterize the electronic properties of ErAs nanoparticles embedded in GaAs pn junctions grown by molecular beam epitaxy. Voltage-dependent capacitance images reveal localized variations in subsurface electronic structure near buried ErAs nanoparticles at lateral length scales of 20-30 nm. Numerical modeling indicates that these variations arise from inhomogeneities in charge modulation due to Fermi level pinning behavior associated with the embedded ErAs nanoparticles. Statistical analysis of image data yields an average particle radius of 6-8 nm—well below the direct resolution limit in scanning capacitance microscopy but discernible via analysis of patterns in nanoscale capacitance images.

Analysis of the Transition in Deformation Mechanisms in Superplastic 5083 Aluminum Alloys by Orientation Imaging Microscopy

DTIC Science & Technology

2001-09-01

Analysis of the Transition in Deformation Mechanisms in Superplastic 5083 Aluminum Alloys by Orientation Imaging Microscopy 6. AUTHOR( S ) Harrell...James W. 5. FUNDING NUMBERS 7. PERFORMING ORGANIZATION NAME( S ) AND ADDRESS(ES) Naval Postgraduate School Monterey, CA 93943-5000 8. PERFORMING...ORGANIZATION REPORT NUMBER 9. SPONSORING / MONITORING AGENCY NAME( S ) AND ADDRESS(ES) General Motors Corp., Research and Development Center

Analysis of Fractured Teeth Utilizing Digital Microscopy: A Pilot Study

DTIC Science & Technology

2016-06-01

ANALYSIS OF FRACTURED TEETH UTILIZING DIGITAL MICROSCOPY: A PILOT STUDY by Thomas Gene Cooper, D.M.D., M.P.H. Lieutenant Commander, Dental Corps...United States Navy A thesis submitted to the Faculty of the Endodontic Graduate Program Naval Postgraduate Dental School Uniformed Services...Postgraduate Dental School Uniformed Services University of the Health Sciences Bethesda, Maryland CERTIFICATE OF APPROVAL MASTER’S THESIS This is to

Polymeric spatial resolution test patterns for mass spectrometry imaging using nano-thermal analysis with atomic force microscopy

DOE PAGES

Tai, Tamin; Kertesz, Vilmos; Lin, Ming -Wei; ...

2017-05-11

As the spatial resolution of mass spectrometry imaging technologies has begun to reach into the nanometer regime, finding readily available or easily made resolution reference materials has become particularly challenging for molecular imaging purposes. This study describes the fabrication, characterization and use of vertical line array polymeric spatial resolution test patterns for nano-thermal analysis/atomic force microscopy/mass spectrometry chemical imaging.

Polymeric spatial resolution test patterns for mass spectrometry imaging using nano-thermal analysis with atomic force microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tai, Tamin; Kertesz, Vilmos; Lin, Ming -Wei

As the spatial resolution of mass spectrometry imaging technologies has begun to reach into the nanometer regime, finding readily available or easily made resolution reference materials has become particularly challenging for molecular imaging purposes. This study describes the fabrication, characterization and use of vertical line array polymeric spatial resolution test patterns for nano-thermal analysis/atomic force microscopy/mass spectrometry chemical imaging.

Detection of local chemical states of lithium and their spatial mapping by scanning transmission electron microscopy, electron energy-loss spectroscopy and hyperspectral image analysis.

PubMed

Muto, Shunsuke; Tatsumi, Kazuyoshi

2017-02-08

Advancements in the field of renewable energy resources have led to a growing demand for the analysis of light elements at the nanometer scale. Detection of lithium is one of the key issues to be resolved for providing guiding principles for the synthesis of cathode active materials, and degradation analysis after repeated use of those materials. We have reviewed the different techniques currently used for the characterization of light elements such as high-resolution transmission electron microscopy, scanning transmission electron microscopy (STEM) and electron energy-loss spectroscopy (EELS). In the present study, we have introduced a methodology to detect lithium in solid materials, particularly for cathode active materials used in lithium-ion battery. The chemical states of lithium were isolated and analyzed from the overlapping multiple spectral profiles, using a suite of STEM, EELS and hyperspectral image analysis. The method was successfully applied in the chemical state analyses of hetero-phases near the surface and grain boundary regions of the active material particles formed by chemical reactions between the electrolyte and the active materials. © The Author 2016. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Clinical laboratory urine analysis: comparison of the UriSed automated microscopic analyzer and the manual microscopy.

PubMed

Ma, Junlong; Wang, Chengbin; Yue, Jiaxin; Li, Mianyang; Zhang, Hongrui; Ma, Xiaojing; Li, Xincui; Xue, Dandan; Qing, Xiaoyan; Wang, Shengjiang; Xiang, Daijun; Cong, Yulong

2013-01-01

Several automated urine sediment analyzers have been introduced to clinical laboratories. Automated microscopic pattern recognition is a new technique for urine particle analysis. We evaluated the analytical and diagnostic performance of the UriSed automated microscopic analyzer and compared with manual microscopy for urine sediment analysis. Precision, linearity, carry-over, and method comparison were carried out. A total of 600 urine samples sent for urinalysis were assessed using the UriSed automated microscopic analyzer and manual microscopy. Within-run and between-run precision of the UriSed for red blood cells (RBC) and white blood cells (WBC) were acceptable at all levels (CV < 20%). Within-run and between-run imprecision of the UriSed testing for cast, squamous epithelial cells (EPI), and bacteria (BAC) were good at middle level and high level (CV < 20%). The linearity analysis revealed substantial agreement between the measured value and the theoretical value of the UriSed for RBC, WBC, cast, EPI, and BAC (r > 0.95). There was no carry-over. RBC, WBC, and squamous epithelial cells with sensitivities and specificities were more than 80% in this study. There is substantial agreement between the UriSed automated microscopic analyzer and the manual microscopy methods. The UriSed provides for a rapid turnaround time.

Isolation and characterization of nanocrystalline cellulose from roselle-derived microcrystalline cellulose.

PubMed

Kian, Lau Kia; Jawaid, Mohammad; Ariffin, Hidayah; Karim, Zoheb

2018-07-15

Roselle fiber is a renewable and sustainable agricultural waste enriched with cellulose polysaccharides. The isolation of Nanocrystalline cellulose (NCC) from roselle-derived microcrystalline cellulose (MCC) is an alternative approach to recover the agricultural roselle plant residue. In the present study, acid hydrolysis with different reaction time was carried out to degrade the roselle-derived MCC to form NCC. The characterizations of isolated NCC were conducted through Fourier Transform Infrared Ray (FTIR), Transmission Electron Microscopy (TEM), Field Emission Scanning Electron Microscopy (FESEM), Atomic Force Microscopy (AFM), Dynamic Light Scattering (DLS), Energy Dispersive Spectroscopy (EDS), X-ray Diffraction (XRD), Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry (DSC). As evaluated from the performed morphological investigations, the needle-like shape NCC nanostructures were observed under TEM and AFM microscopy studies, while irregular rod-like shape of NCC was observed under FESEM analysis. With 60min hydrolysis time, XRD analysis demonstrated the highest NCC crystallinity degree with 79.5%. In thermal analysis by TGA and DSC, the shorter hydrolysis time tended to produce NCC with higher thermal stability. Thus, the isolated NCC from roselle-derived MCC has high potential to be used in application of pharmaceutical and biomedical fields for nanocomposite fabrication. Copyright © 2018 Elsevier B.V. All rights reserved.

In vivo chemical and structural analysis of plant cuticular waxes using stimulated Raman scattering microscopy.

PubMed

Littlejohn, George R; Mansfield, Jessica C; Parker, David; Lind, Rob; Perfect, Sarah; Seymour, Mark; Smirnoff, Nicholas; Love, John; Moger, Julian

2015-05-01

The cuticle is a ubiquitous, predominantly waxy layer on the aerial parts of higher plants that fulfils a number of essential physiological roles, including regulating evapotranspiration, light reflection, and heat tolerance, control of development, and providing an essential barrier between the organism and environmental agents such as chemicals or some pathogens. The structure and composition of the cuticle are closely associated but are typically investigated separately using a combination of structural imaging and biochemical analysis of extracted waxes. Recently, techniques that combine stain-free imaging and biochemical analysis, including Fourier transform infrared spectroscopy microscopy and coherent anti-Stokes Raman spectroscopy microscopy, have been used to investigate the cuticle, but the detection sensitivity is severely limited by the background signals from plant pigments. We present a new method for label-free, in vivo structural and biochemical analysis of plant cuticles based on stimulated Raman scattering (SRS) microscopy. As a proof of principle, we used SRS microscopy to analyze the cuticles from a variety of plants at different times in development. We demonstrate that the SRS virtually eliminates the background interference compared with coherent anti-Stokes Raman spectroscopy imaging and results in label-free, chemically specific confocal images of cuticle architecture with simultaneous characterization of cuticle composition. This innovative use of the SRS spectroscopy may find applications in agrochemical research and development or in studies of wax deposition during leaf development and, as such, represents an important step in the study of higher plant cuticles. © 2015 American Society of Plant Biologists. All Rights Reserved.

Strategies for rare-event detection: an approach for automated fetal cell detection in maternal blood.

PubMed Central

Oosterwijk, J C; Knepflé, C F; Mesker, W E; Vrolijk, H; Sloos, W C; Pattenier, H; Ravkin, I; van Ommen, G J; Kanhai, H H; Tanke, H J

1998-01-01

This article explores the feasibility of the use of automated microscopy and image analysis to detect the presence of rare fetal nucleated red blood cells (NRBCs) circulating in maternal blood. The rationales for enrichment and for automated image analysis for "rare-event" detection are reviewed. We also describe the application of automated image analysis to 42 maternal blood samples, using a protocol consisting of one-step enrichment followed by immunocytochemical staining for fetal hemoglobin (HbF) and FISH for X- and Y-chromosomal sequences. Automated image analysis consisted of multimode microscopy and subsequent visual evaluation of image memories containing the selected objects. The FISH results were compared with the results of conventional karyotyping of the chorionic villi. By use of manual screening, 43% of the slides were found to be positive (>=1 NRBC), with a mean number of 11 NRBCs (range 1-40). By automated microscopy, 52% were positive, with on average 17 NRBCs (range 1-111). There was a good correlation between both manual and automated screening, but the NRBC yield from automated image analysis was found to be superior to that from manual screening (P=.0443), particularly when the NRBC count was >15. Seven (64%) of 11 XY fetuses were correctly diagnosed by FISH analysis of automatically detected cells, and all discrepancies were restricted to the lower cell-count range. We believe that automated microscopy and image analysis reduce the screening workload, are more sensitive than manual evaluation, and can be used to detect rare HbF-containing NRBCs in maternal blood. PMID:9837832

Microfluidics and Raman microscopy: current applications and future challenges.

PubMed

Chrimes, Adam F; Khoshmanesh, Khashayar; Stoddart, Paul R; Mitchell, Arnan; Kalantar-Zadeh, Kourosh

2013-07-07

Raman microscopy systems are becoming increasingly widespread and accessible for characterising chemical species. Microfluidic systems are also progressively finding their way into real world applications. Therefore, it is anticipated that the integration of Raman systems with microfluidics will become increasingly attractive and practical. This review aims to provide an overview of Raman microscopy-microfluidics integrated systems for researchers who are actively interested in utilising these tools. The fundamental principles and application strengths of Raman microscopy are discussed in the context of microfluidics. Various configurations of microfluidics that incorporate Raman microscopy methods are presented, with applications highlighted. Data analysis methods are discussed, with a focus on assisting the interpretation of Raman-microfluidics data from complex samples. Finally, possible future directions of Raman-microfluidic systems are presented.

Insights into the prominent effect of mahanimbine on Acanthamoeba castellanii: Cell profiling analysis based on microscopy techniques

NASA Astrophysics Data System (ADS)

Hashim, Fatimah; Amin, Nakisah Mat

2017-02-01

Mahanimbine (MH), has been shown to have antiamoeba properties. Therefore, the aim of this study was to assess the growth inhibitory mechanisms of MH on Acanthamoeba castellanii, a causative agents for Acanthamoeba keratitis. The IC50 value obtained for MH against A. castellanii was 1.18 µg/ml. Light and scanning electron microscopy observation showed that most cells were in cystic appearance. While transmission electron microscopy observation revealed changes at the ultrastructural level and fluorescence microscopy observation indicated the induction of apoptosis and autophagic activity in the amoeba cytoplasms. In conclusion, MH has very potent anti-amoebic properties on A. castellanii as is shown by cytotoxicity analyses based on microscopy techniques.

Ultrathin inorganic molecular nanowire based on polyoxometalates

PubMed Central

Zhang, Zhenxin; Murayama, Toru; Sadakane, Masahiro; Ariga, Hiroko; Yasuda, Nobuhiro; Sakaguchi, Norihito; Asakura, Kiyotaka; Ueda, Wataru

2015-01-01

The development of metal oxide-based molecular wires is important for fundamental research and potential practical applications. However, examples of these materials are rare. Here we report an all-inorganic transition metal oxide molecular wire prepared by disassembly of larger crystals. The wires are comprised of molybdenum(VI) with either tellurium(IV) or selenium(IV): {(NH4)2[XMo6O21]}n (X=tellurium(IV) or selenium(IV)). The ultrathin molecular nanowires with widths of 1.2 nm grow to micrometre-scale crystals and are characterized by single-crystal X-ray analysis, Rietveld analysis, scanning electron microscopy, X-ray photoelectron spectroscopy, ultraviolet–visible spectroscopy, thermal analysis and elemental analysis. The crystals can be disassembled into individual molecular wires through cation exchange and subsequent ultrasound treatment, as visualized by atomic force microscopy and transmission electron microscopy. The ultrathin molecular wire-based material exhibits high activity as an acid catalyst, and the band gap of the molecular wire-based crystal is tunable by heat treatment. PMID:26139011

Carbon Nanostructure Examined by Lattice Fringe Analysis of High Resolution Transmission Electron Microscopy Images

NASA Technical Reports Server (NTRS)

VanderWal, Randy L.; Tomasek, Aaron J.; Street, Kenneth; Thompson, William K.

2002-01-01

The dimensions of graphitic layer planes directly affect the reactivity of soot towards oxidation and growth. Quantification of graphitic structure could be used to develop and test correlations between the soot nanostructure and its reactivity. Based upon transmission electron microscopy images, this paper provides a demonstration of the robustness of a fringe image analysis code for determining the level of graphitic structure within nanoscale carbon, i.e. soot. Results, in the form of histograms of graphitic layer plane lengths, are compared to their determination through Raman analysis.

Carbon Nanostructure Examined by Lattice Fringe Analysis of High Resolution Transmission Electron Microscopy Images

NASA Technical Reports Server (NTRS)

VanderWal, Randy L.; Tomasek, Aaron J.; Street, Kenneth; Thompson, William K.; Hull, David R.

2003-01-01

The dimensions of graphitic layer planes directly affect the reactivity of soot towards oxidation and growth. Quantification of graphitic structure could be used to develop and test correlations between the soot nanostructure and its reactivity. Based upon transmission electron microscopy images, this paper provides a demonstration of the robustness of a fringe image analysis code for determining the level of graphitic structure within nanoscale carbon, i.e., soot. Results, in the form of histograms of graphitic layer plane lengths, are compared to their determination through Raman analysis.

STED super-resolution microscopy of clinical paraffin-embedded human rectal cancer tissue.

PubMed

Ilgen, Peter; Stoldt, Stefan; Conradi, Lena-Christin; Wurm, Christian Andreas; Rüschoff, Josef; Ghadimi, B Michael; Liersch, Torsten; Jakobs, Stefan

2014-01-01

Formalin fixed and paraffin-embedded human tissue resected during cancer surgery is indispensable for diagnostic and therapeutic purposes and represents a vast and largely unexploited resource for research. Optical microscopy of such specimen is curtailed by the diffraction-limited resolution of conventional optical microscopy. To overcome this limitation, we used STED super-resolution microscopy enabling optical resolution well below the diffraction barrier. We visualized nanoscale protein distributions in sections of well-annotated paraffin-embedded human rectal cancer tissue stored in a clinical repository. Using antisera against several mitochondrial proteins, STED microscopy revealed distinct sub-mitochondrial protein distributions, suggesting a high level of structural preservation. Analysis of human tissues stored for up to 17 years demonstrated that these samples were still amenable for super-resolution microscopy. STED microscopy of sections of HER2 positive rectal adenocarcinoma revealed details in the surface and intracellular HER2 distribution that were blurred in the corresponding conventional images, demonstrating the potential of super-resolution microscopy to explore the thus far largely untapped nanoscale regime in tissues stored in biorepositories.

STED Super-Resolution Microscopy of Clinical Paraffin-Embedded Human Rectal Cancer Tissue

PubMed Central

Wurm, Christian Andreas; Rüschoff, Josef; Ghadimi, B. Michael; Liersch, Torsten; Jakobs, Stefan

2014-01-01

Formalin fixed and paraffin-embedded human tissue resected during cancer surgery is indispensable for diagnostic and therapeutic purposes and represents a vast and largely unexploited resource for research. Optical microscopy of such specimen is curtailed by the diffraction-limited resolution of conventional optical microscopy. To overcome this limitation, we used STED super-resolution microscopy enabling optical resolution well below the diffraction barrier. We visualized nanoscale protein distributions in sections of well-annotated paraffin-embedded human rectal cancer tissue stored in a clinical repository. Using antisera against several mitochondrial proteins, STED microscopy revealed distinct sub-mitochondrial protein distributions, suggesting a high level of structural preservation. Analysis of human tissues stored for up to 17 years demonstrated that these samples were still amenable for super-resolution microscopy. STED microscopy of sections of HER2 positive rectal adenocarcinoma revealed details in the surface and intracellular HER2 distribution that were blurred in the corresponding conventional images, demonstrating the potential of super-resolution microscopy to explore the thus far largely untapped nanoscale regime in tissues stored in biorepositories. PMID:25025184

Identification of nodal tissue in the living heart using rapid scanning fiber-optics confocal microscopy and extracellular fluorophores.

PubMed

Huang, Chao; Kaza, Aditya K; Hitchcock, Robert W; Sachse, Frank B

2013-09-01

Risks associated with pediatric reconstructive heart surgery include injury of the sinoatrial node (SAN) and atrioventricular node (AVN), requiring cardiac rhythm management using implantable pacemakers. These injuries are the result of difficulties in identifying nodal tissues intraoperatively. Here we describe an approach based on confocal microscopy and extracellular fluorophores to quantify tissue microstructure and identify nodal tissue. Using conventional 3-dimensional confocal microscopy we investigated the microstructural arrangement of SAN, AVN, and atrial working myocardium (AWM) in fixed rat heart. AWM exhibited a regular striated arrangement of the extracellular space. In contrast, SAN and AVN had an irregular, reticulated arrangement. AWM, SAN, and AVN tissues were beneath a thin surface layer of tissue that did not obstruct confocal microscopic imaging. Subsequently, we imaged tissues in living rat hearts with real-time fiber-optics confocal microscopy. Fiber-optics confocal microscopy images resembled images acquired with conventional confocal microscopy. We investigated spatial regularity of tissue microstructure from Fourier analysis and second-order image moments. Fourier analysis of fiber-optics confocal microscopy images showed that the spatial regularity of AWM was greater than that of nodal tissues (37.5 ± 5.0% versus 24.3 ± 3.9% for SAN and 23.8 ± 3.7% for AVN; P<0.05). Similar differences of spatial regularities were revealed from second-order image moments (50.0 ± 7.3% for AWM versus 29.3 ± 6.7% for SAN and 27.3 ± 5.5% for AVN; P<0.05). The study demonstrates feasibility of identifying nodal tissue in living heart using extracellular fluorophores and fiber-optics confocal microscopy. Application of the approach in pediatric reconstructive heart surgery may reduce risks of injuring nodal tissues.

Potential of ultraviolet wide-field imaging and multiphoton microscopy for analysis of dehydroergosterol in cellular membranes.

PubMed

Wüstner, Daniel; Brewer, Jonathan R; Bagatolli, Luis; Sage, Daniel

2011-01-01

Dehydroergosterol (DHE) is an intrinsically fluorescent sterol with absorption/emission in the ultraviolet (UV) region and biophysical properties similar to those of cholesterol. We compared the potential of UV-sensitive low-light-level wide-field (UV-WF) imaging with that of multiphoton (MP) excitation microscopy to monitor DHE in living cells. Significantly reduced photobleaching in MP microscopy of DHE enabled us to acquire three-dimensional z-stacks of DHE-stained cells and to obtain high-resolution maps of DHE in surface ruffles, nanotubes, and the apical membrane of epithelial cells. We found that the lateral resolution of MP microscopy is ∼1.5-fold higher than that of UV-WF deconvolution microscopy, allowing for improved spatiotemporal analysis of plasma membrane sterol distribution. Surface intensity patterns of DHE with a diameter of 0.2 μm persisting over several minutes could be resolved by MP time-lapse microscopy. Diffusion coefficients of 0.25-μm-diameter endocytic vesicles containing DHE were determined by MP spatiotemporal image correlation spectroscopy. The requirement of extremely high laser power for visualization of DHE by MP microscopy made this method less potent for multicolor applications with organelle markers like green fluorescent protein-tagged proteins. The signal-to-noise ratio obtainable by UV-WF imaging could be significantly improved by pixelwise bleach rate fitting and calculation of an amplitude image from the decay model and by frame averaging after pixelwise bleaching correction of the image stacks. We conclude that UV-WF imaging and MP microscopy of DHE provide complementary information regarding membrane distribution and intracellular targeting of sterols. © 2010 Wiley-Liss, Inc.

Dark Field Microscopy for Analytical Laboratory Courses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Augspurger, Ashley E; Stender, Anthony S; Marchuk, Kyle

2014-06-10

An innovative and inexpensive optical microscopy experiment for a quantitative analysis or an instrumental analysis chemistry course is described. The students have hands-on experience with a dark field microscope and investigate the wavelength dependence of localized surface plasmon resonance in gold and silver nanoparticles. Students also observe and measure individual crystal growth during a replacement reaction between copper and silver nitrate. The experiment allows for quantitative, qualitative, and image data analyses for undergraduate students.

A flow-cytometry-based method to simplify the analysis and quantification of protein association to chromatin in mammalian cells

PubMed Central

Forment, Josep V.; Jackson, Stephen P.

2016-01-01

Protein accumulation on chromatin has traditionally been studied using immunofluorescence microscopy or biochemical cellular fractionation followed by western immunoblot analysis. As a way to improve the reproducibility of this kind of analysis, make it easier to quantify and allow a stream-lined application in high-throughput screens, we recently combined a classical immunofluorescence microscopy detection technique with flow cytometry1. In addition to the features described above, and by combining it with detection of both DNA content and DNA replication, this method allows unequivocal and direct assignment of cell-cycle distribution of protein association to chromatin without the need for cell culture synchronization. Furthermore, it is relatively quick (no more than a working day from sample collection to quantification), requires less starting material compared to standard biochemical fractionation methods and overcomes the need for flat, adherent cell types that are required for immunofluorescence microscopy. PMID:26226461

Analysis and modification of defective surface aggregates on PCDTBT:PCBM solar cell blends using combined Kelvin probe, conductive and bimodal atomic force microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Noh, Hanaul; Diaz, Alfredo J.; Solares, Santiago D.

Organic photovoltaic systems comprising donor polymers and acceptor fullerene derivatives are attractive for inexpensive energy harvesting. Extensive research on polymer solar cells has provided insight into the factors governing device-level efficiency and stability. However, the detailed investigation of nanoscale structures is still challenging. Here we demonstrate the analysis and modification of unidentified surface aggregates. The aggregates are characterized electrically by Kelvin probe force microscopy and conductive atomic force microscopy (C-AFM), whereby the correlation between local electrical potential and current confirms a defective charge transport. Bimodal AFM modification confirms that the aggregates exist on top of the solar cell structure, andmore » is used to remove them and to reveal the underlying active layer. The systematic analysis of the surface aggregates suggests that the structure consists of PCBM molecules.« less

The quantitative analysis of silicon carbide surface smoothing by Ar and Xe cluster ions

NASA Astrophysics Data System (ADS)

Ieshkin, A. E.; Kireev, D. S.; Ermakov, Yu. A.; Trifonov, A. S.; Presnov, D. E.; Garshev, A. V.; Anufriev, Yu. V.; Prokhorova, I. G.; Krupenin, V. A.; Chernysh, V. S.

2018-04-01

The gas cluster ion beam technique was used for the silicon carbide crystal surface smoothing. The effect of processing by two inert cluster ions, argon and xenon, was quantitatively compared. While argon is a standard element for GCIB, results for xenon clusters were not reported yet. Scanning probe microscopy and high resolution transmission electron microscopy techniques were used for the analysis of the surface roughness and surface crystal layer quality. The gas cluster ion beam processing results in surface relief smoothing down to average roughness about 1 nm for both elements. It was shown that xenon as the working gas is more effective: sputtering rate for xenon clusters is 2.5 times higher than for argon at the same beam energy. High resolution transmission electron microscopy analysis of the surface defect layer gives values of 7 ± 2 nm and 8 ± 2 nm for treatment with argon and xenon clusters.

Analysis and modification of defective surface aggregates on PCDTBT:PCBM solar cell blends using combined Kelvin probe, conductive and bimodal atomic force microscopy

DOE PAGES

Noh, Hanaul; Diaz, Alfredo J.; Solares, Santiago D.

2017-03-08

Organic photovoltaic systems comprising donor polymers and acceptor fullerene derivatives are attractive for inexpensive energy harvesting. Extensive research on polymer solar cells has provided insight into the factors governing device-level efficiency and stability. However, the detailed investigation of nanoscale structures is still challenging. Here we demonstrate the analysis and modification of unidentified surface aggregates. The aggregates are characterized electrically by Kelvin probe force microscopy and conductive atomic force microscopy (C-AFM), whereby the correlation between local electrical potential and current confirms a defective charge transport. Bimodal AFM modification confirms that the aggregates exist on top of the solar cell structure, andmore » is used to remove them and to reveal the underlying active layer. The systematic analysis of the surface aggregates suggests that the structure consists of PCBM molecules.« less

Ceramic femoral component fracture in total knee arthroplasty: an analysis using fractography, fourier-transform infrared microscopy, contact radiography and histology.

PubMed

Krueger, Alexander P; Singh, Gurpal; Beil, Frank Timo; Feuerstein, Bernd; Ruether, Wolfgang; Lohmann, Christoph H

2014-05-01

Ceramic components in total knee arthroplasty (TKA) are evolving. We analyze the first case of BIOLOX delta ceramic femoral component fracture. A longitudinal midline fracture in the patellar groove was present, with an intact cement mantle and no bony defects. Fractographic analysis with laser scanning microscopy and white light interferometry showed no evidence of arrest lines, hackles, wake hackles, material flaws, fatigue or crack propagation. Analysis of periprosthetic tissues with Fourier-transform infrared (FT-IR) microscopy, contact radiography, histology, and subsequent digestion and high-speed centrifugation did not show ceramic debris. A macrophage-dominated response was present around polyethylene debris. We conclude that ceramic femoral component failure in this case was related to a traumatic event. Further research is needed to determine the suitability of ceramic components in TKA. Copyright © 2014 Elsevier Inc. All rights reserved.

Analysis and modification of defective surface aggregates on PCDTBT:PCBM solar cell blends using combined Kelvin probe, conductive and bimodal atomic force microscopy

PubMed Central

Noh, Hanaul; Diaz, Alfredo J

2017-01-01

Organic photovoltaic systems comprising donor polymers and acceptor fullerene derivatives are attractive for inexpensive energy harvesting. Extensive research on polymer solar cells has provided insight into the factors governing device-level efficiency and stability. However, the detailed investigation of nanoscale structures is still challenging. Here we demonstrate the analysis and modification of unidentified surface aggregates. The aggregates are characterized electrically by Kelvin probe force microscopy and conductive atomic force microscopy (C-AFM), whereby the correlation between local electrical potential and current confirms a defective charge transport. Bimodal AFM modification confirms that the aggregates exist on top of the solar cell structure, and is used to remove them and to reveal the underlying active layer. The systematic analysis of the surface aggregates suggests that the structure consists of PCBM molecules. PMID:28382247

Analysis of multi-channel microscopy: Spectral self-interference, multi-detector confocal and 4Pi systems

NASA Astrophysics Data System (ADS)

Davis, Brynmor J.

Fluorescence microscopy is an important and ubiquitous tool in biological imaging due to the high specificity with which fluorescent molecules can be attached to an organism and the subsequent nondestructive in-vivo imaging allowed. Focused-light microscopies allow three-dimensional fluorescence imaging but their resolution is restricted by diffraction. This effect is particularly limiting in the axial dimension as the diffraction-limited focal volume produced by a lens is more extensive along the optical axis than perpendicular to it. Approaches such as confocal microscopy and 4Pi microscopy have been developed to improve the axial resolution. Spectral Self-Interference Fluorescence Microscopy (SSFM) is another high-axial-resolution technique and is the principal subject of this dissertation. Nanometer-precision localization of a single fluorescent layer has been demonstrated using SSFM. This accuracy compares favorably with the axial resolutions given by confocal and 4Pi systems at similar operating parameters (these resolutions are approximately 350nm and 80nm respectively). This theoretical work analyzes the expected performance of the SSFM system when imaging a general object, i.e. an arbitrary fluorophore density function rather than a single layer. An existing model of SSFM is used in simulations to characterize the system's resolution. Several statistically-based reconstruction methods are applied to show that the expected resolution for SSFM is similar to 4Pi microscopy for a general object but does give very high localization accuracy when the object is known to consist of a limited number of layers. SSFM is then analyzed in a linear systems framework and shown to have strong connections, both physically and mathematically, to a multi-channel 4Pi microscope. Fourier-domain analysis confirms that SSFM cannot be expected to outperform this multi-channel 4Pi instrument. Differences between the channels in spatial-scanning, multi-channel microscopies are then exploited to show that such instruments can operate at a sub-Nyquist scanning rate but still produce images largely free of aliasing effects. Multi-channel analysis is also used to show how light typically discarded in confocal and 4Pi systems can be collected and usefully incorporated into the measured image.

In vivo non-invasive monitoring of collagen remodelling by two-photon microscopy after micro-ablative fractional laser resurfacing.

PubMed

Cicchi, Riccardo; Kapsokalyvas, Dimitrios; Troiano, Michela; Campolmi, Piero; Morini, Cristiano; Massi, Daniela; Cannarozzo, Giovanni; Lotti, Torello; Pavone, Francesco Saverio

2014-11-01

Non-linear optical microscopy is becoming popular as a non-invasive in vivo imaging modality in dermatology. In this study, combined TPF and SHG microscopy were used to monitor collagen remodelling in vivo after micro-ablative fractional laser resurfacing. Papillary dermis of living subjects, covering a wide age range, was imaged immediately before and forty days after treatment. A qualitative visual examination of acquired images demonstrated an age-dependent remodelling effect on collagen. Additional quantitative analysis of new collagen production was performed by means of two image analysis methods. A higher increase in SHG to TPF ratio, corresponding to a stronger treatment effectiveness, was found in older subjects, whereas the effect was found to be negligible in young, and minimal in middle age subjects. Analysis of collagen images also showed a dependence of the treatment effectiveness with age but with controversial results. While the diagnostic potential of in vivo multiphoton microscopy has already been demonstrated for skin cancer and other skin diseases, here we first successfully explore its potential use for a non-invasive follow-up of a laser-based treatment. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Combined use of confocal laser scanning microscopy (CLSM) and Raman microscopy (RM): investigations on EPS-Matrix.

PubMed

Wagner, Michael; Ivleva, Natalia P; Haisch, Christoph; Niessner, Reinhard; Horn, Harald

2009-01-01

Confocal laser scanning microscopy (CLSM) was applied in combination with Raman microscopy (RM) for the characterization of heterotrophic biofilms. Compared to CLSM, RM allows for a deeper insight into the chemical structure of extracellular polymeric substances (EPS) of the biofilm matrix. A low load of glucose (2 g m(-2)d(-1)) was applied as substrate to ensure small growth rates of the heterotrophic biofilm. To investigate the influence of hydrodynamic conditions on the chemical composition of EPS, a three funnel flow system was used, wherein biofilms were grown at Reynolds numbers of 1000, 2500 and 4000, respectively. 31 and 92 days after inoculation with activated sludge supernatant RM was applied as an additional technique to standard CLSM measurements for a more detailed analysis of the biofilm matrix. Polysaccharide-related Raman bands are in good agreement with the lectin binding analysis from CLSM. For the older biofilm, lectin binding analysis showed no change in the composition of EPS, whereas Raman spectra pointed out a change of EPS composition from predominantly polysaccharides to predominantly (glyco) proteins. For the applied substrate condition no significant influence of the Reynolds number on the chemical properties was observed.

Nuclear microscopy in biomedical analysis with special emphasis on clinical metal biology

NASA Astrophysics Data System (ADS)

Lindh, Ulf; Frisk, Peter; Nyström, Joakim; Danersund, Antero; Hudecek, Romuald; Lindvall, Anders; Thunell, Stig

1997-07-01

Nuclear microscopy based upon developments in high energy ion beam techniques is by now an accepted technique in many fields of research. The advancements into the biomedical field have, however, been slower than expected. A major factor explaining this tendency is the availability of nuclear microscopy. This paper reviews briefly the biomedical work using nuclear microscopy that has been carried out since the 4 th International Conference on Nuclear Microprobe Technology and Applications held in Shanghai. Nuclear microscopy of isolated individual blood cells from patients adversely affected by metal exposure from dental amalgam has been performed both before and after removal of the metallic fillings. The elemental profile of blood cells was more or less normalised after treatment. Some of these results will be presented to illustrate a medical application. Results from bulk analysis by ICP-MS of erythrocytes and plasma before and after treatment will also be presented to illustrate the difference in information content between these two approaches as well as the need for complementary information in solving biomedical problems. As part of a larger study of acute porphyria, nuclear microscopy of blood cells was included among the 78 laboratory tests. The approach in this study was unbiased in the sense that no hypothesis was formulated as to which laboratory parameters would be the most explanatory for health or disease. Multivariate discriminant analysis was applied to the large amounts of data acquired. This approach led to the hypothesis that oxidative stress increased the synthesis of manganese-dependent superoxide dismutase in the mitochondria of polymorphonuclear leukocytes, explaining the increase of manganese in these cells. Antioxidant therapy was therefore applied to a couple of patients with porphyria, however, without clinical success.

STATISTICAL ANALYSIS OF SPECTROPHOTOMETRIC DETERMINATIONS OF BORON; Estudo Estatistico de Determinacoes Espectrofotometricas de Boro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lima, F.W.; Pagano, C.; Schneiderman, B.

1959-07-01

Boron can be determined quantitatively by absorption spectrophotometry of solutions of the red compound formed by the reaction of boric acid with curcumin. This reaction is affected by various factors, some of which can be detected easily in the data interpretation. Others, however, provide more difficulty. The application of modern statistical method to the study of the influence of these factors on the quantitative determination of boron is presented. These methods provide objective ways of establishing significant effects of the factors involved. (auth)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Phuthong, Witchukorn; Huang, Zubin; Wittkopp, Tyler M.

To investigate the dynamics of photosynthetic pigment-protein complexes in vascular plants at high resolution in an aqueous environment, membrane-protruding oxygen-evolving complexes (OECs) associated with photosystem II (PSII) on spinach ( Spinacia oleracea) grana membranes were examined using contact mode atomic force microscopy. This study represents, to our knowledge, the first use of atomic force microscopy to distinguish the putative large extrinsic loop of Photosystem II CP47 reaction center protein (CP47) from the putative oxygen-evolving enhancer proteins 1, 2, and 3 (PsbO, PsbP, and PsbQ) and large extrinsic loop of Photosystem II CP43 reaction center protein (CP43) in the PSII-OEC extrinsicmore » domains of grana membranes under conditions resulting in the disordered arrangement of PSII-OEC particles. Moreover, we observed uncharacterized membrane particles that, based on their physical characteristics and electrophoretic analysis of the polypeptides associated with the grana samples, are hypothesized to be a domain of photosystem I that protrudes from the stromal face of single thylakoid bilayers. Furthermore, our results are interpreted in the context of the results of others that were obtained using cryo-electron microscopy (and single particle analysis), negative staining and freeze-fracture electron microscopy, as well as previous atomic force microscopy studies.« less

Kinetic Modeling of Accelerated Stability Testing Enabled by Second Harmonic Generation Microscopy.

PubMed

Song, Zhengtian; Sarkar, Sreya; Vogt, Andrew D; Danzer, Gerald D; Smith, Casey J; Gualtieri, Ellen J; Simpson, Garth J

2018-04-03

The low limits of detection afforded by second harmonic generation (SHG) microscopy coupled with image analysis algorithms enabled quantitative modeling of the temperature-dependent crystallization of active pharmaceutical ingredients (APIs) within amorphous solid dispersions (ASDs). ASDs, in which an API is maintained in an amorphous state within a polymer matrix, are finding increasing use to address solubility limitations of small-molecule APIs. Extensive stability testing is typically performed for ASD characterization, the time frame for which is often dictated by the earliest detectable onset of crystal formation. Here a study of accelerated stability testing on ritonavir, a human immunodeficiency virus (HIV) protease inhibitor, has been conducted. Under the condition for accelerated stability testing at 50 °C/75%RH and 40 °C/75%RH, ritonavir crystallization kinetics from amorphous solid dispersions were monitored by SHG microscopy. SHG microscopy coupled by image analysis yielded limits of detection for ritonavir crystals as low as 10 ppm, which is about 2 orders of magnitude lower than other methods currently available for crystallinity detection in ASDs. The four decade dynamic range of SHG microscopy enabled quantitative modeling with an established (JMAK) kinetic model. From the SHG images, nucleation and crystal growth rates were independently determined.

Principal component and spatial correlation analysis of spectroscopic-imaging data in scanning probe microscopy.

PubMed

Jesse, Stephen; Kalinin, Sergei V

2009-02-25

An approach for the analysis of multi-dimensional, spectroscopic-imaging data based on principal component analysis (PCA) is explored. PCA selects and ranks relevant response components based on variance within the data. It is shown that for examples with small relative variations between spectra, the first few PCA components closely coincide with results obtained using model fitting, and this is achieved at rates approximately four orders of magnitude faster. For cases with strong response variations, PCA allows an effective approach to rapidly process, de-noise, and compress data. The prospects for PCA combined with correlation function analysis of component maps as a universal tool for data analysis and representation in microscopy are discussed.

Multiplex real-time quantitative PCR, microscopy and rapid diagnostic immuno-chromatographic tests for the detection of Plasmodium spp: performance, limit of detection analysis and quality assurance.

PubMed

Khairnar, Krishna; Martin, Donald; Lau, Rachel; Ralevski, Filip; Pillai, Dylan R

2009-12-09

Accurate laboratory diagnosis of malaria species in returning travelers is paramount in the treatment of this potentially fatal infectious disease. A total of 466 blood specimens from returning travelers to Africa, Asia, and South/Central America with suspected malaria infection were collected between 2007 and 2009 at the reference public health laboratory. These specimens were assessed by reference microscopy, multipex real-time quantitative polymerase chain reaction (QPCR), and two rapid diagnostic immuno-chromatographic tests (ICT) in a blinded manner. Key clinical laboratory parameters such as limit of detection (LOD) analysis on clinical specimens by parasite stage, inter-reader variability of ICTs, staffing implications, quality assurance and cost analysis were evaluated. QPCR is the most analytically sensitive method (sensitivity 99.41%), followed by CARESTART (sensitivity 88.24%), and BINAXNOW (sensitivity 86.47%) for the diagnosis of malaria in returning travelers when compared to reference microscopy. However, microscopy was unable to specifically identify Plasmodia spp. in 18 out of 170 positive samples by QPCR. Moreover, the 17 samples that were negative by microscopy and positive by QPCR were also positive by ICTs. Quality assurance was achieved for QPCR by exchanging a blinded proficiency panel with another reference laboratory. The Kappa value of inter-reader variability among three readers for BINAXNOW and CARESTART was calculated to be 0.872 and 0.898 respectively. Serial dilution studies demonstrated that the QPCR cycle threshold correlates linearly with parasitemia (R(2) = 0.9746) in a clinically relevant dynamic range and retains a LOD of 11 rDNA copies/microl for P. falciparum, which was several log lower than reference microscopy and ICTs. LOD for QPCR is affected not only by parasitemia but the parasite stage distribution of each clinical specimen. QPCR was approximately 6-fold more costly than reference microscopy. These data suggest that multiplex QPCR although more costly confers a significant diagnostic advantage in terms of LOD compared to reference microscopy and ICTs for all four species. Quality assurance of QPCR is essential to the maintenance of proficiency in the clinical laboratory. ICTs showed good concordance between readers however lacked sensitivity for non-falciparum species due to antigenic differences and low parasitemia. Multiplex QPCR but not ICTs is an essential adjunct to microscopy in the reference laboratory detection of malaria species specifically due to the superior LOD. ICTs are better suited to the non-reference laboratory where lower specimen volumes challenge microscopy proficiency in the non-endemic setting.

Morphology- and orientation-controlled gallium arsenide nanowires on silicon substrates.

PubMed

Ihn, Soo-Ghang; Song, Jong-In; Kim, Tae-Wook; Leem, Dong-Seok; Lee, Takhee; Lee, Sang-Geul; Koh, Eui Kwan; Song, Kyung

2007-01-01

GaAs nanowires were epitaxially grown on Si(001) and Si(111) substrates by using Au-catalyzed vapor-liquid-solid (VLS) growth in a solid source molecular beam epitaxy system. Scanning electron microscopy analysis revealed that almost all the GaAs nanowires were grown along <111> directions on both Si substrates for growth conditions investigated. The GaAs nanowires had a very uniform diameter along the growth direction. X-ray diffraction data and transmission electron microscopy analysis revealed that the GaAs<111> nanowires had a mixed crystal structure of the hexagonal wurtzite and the cubic zinc-blende. Current-voltage characteristics of junctions formed by the epitaxially grown GaAs nanowires and the Si substrate were investigated by using a current-sensing atomic force microscopy.

Direct observation of anti-phase boundaries in heteroepitaxy of GaSb thin films grown on Si(001) by transmission electron microscopy

NASA Astrophysics Data System (ADS)

Woo, S. Y.; Hosseini Vajargah, S.; Ghanad-Tavakoli, S.; Kleiman, R. N.; Botton, G. A.

2012-10-01

Unambiguous identification of anti-phase boundaries (APBs) in heteroepitaxial films of GaSb grown on Si has been so far elusive. In this work, we present conventional transmission electron microscopy (TEM) diffraction contrast imaging using superlattice reflections, in conjunction with convergent beam electron diffraction analysis, to determine a change in polarity across APBs in order to confirm the presence of anti-phase disorder. In-depth analysis of anti-phase disorder is further supported with atomic resolution high-angle annular dark-field scanning transmission electron microscopy. The nature of APBs in GaSb is further elucidated by a comparison to previous results for GaAs epilayers grown on Si.

Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection.

PubMed

Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H; Nørregaard, Rikke; Møller-Jensen, Jakob; Nejsum, Lene N

2017-10-01

Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacterial colonies in infected host cells (Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy, Ernstsen et al., 2017 [1]). The infected cells were imaged with a 10× objective and number of intracellular bacterial colonies, their size distribution and the number of cell nuclei were automatically quantified using a spot detection-tool. The spot detection-output was exported to Excel, where data analysis was performed. In this article, micrographs and spot detection data are made available to facilitate implementation of the method.

Combining kriging, multispectral and multimodal microscopy to resolve malaria-infected erythrocyte contents.

PubMed

Dabo-Niang, S; Zoueu, J T

2012-09-01

In this communication, we demonstrate how kriging, combine with multispectral and multimodal microscopy can enhance the resolution of malaria-infected images and provide more details on their composition, for analysis and diagnosis. The results of this interpolation applied to the two principal components of multispectral and multimodal images illustrate that the examination of the content of Plasmodium falciparum infected human erythrocyte is improved. © 2012 The Authors Journal of Microscopy © 2012 Royal Microscopical Society.

Scanning probe microscopy in mineralogical studies: about origin of the observed roughness of natural silica-rich glasses

NASA Astrophysics Data System (ADS)

Golubev, Ye A.; Isaenko, S. I.

2017-10-01

We have studied different mineralogical objects: natural glasses of impact (tektites, impactites) and volcanic (obsidians) origin, using atomic force microscopy, X-ray microanalysis, infrared and Raman spectroscopy. The spectroscopy showed the difference in the structure and chemical composition of the glasses of different origin. The analysis of the dependence of nanoscale heterogeneity of the glasses, revealed by the atomic force microscopy, on their structural and chemical features was carried out.

Wirelessly controlled micro- and nanostructures for bioapplications

NASA Astrophysics Data System (ADS)

Lindo, Andre Machado

Apresentar uma contribuicao para a caracterizacao e compreensao do comportamento hidrodinâmico da laguna Ria de Aveiro, baseada num estudo experimental e num estudo de modelacao numerica, e o objectivo deste trabalho. As caracteristicas hidrologicas da Ria de Aveiro foram investigadas atraves da realizacao de varias campanhas de amostragem, tendo sido efectuadas medicoes de altura de agua, salinidade, temperatura da agua e velocidade da corrente em varias estacoes distribuidas ao longo de quatro canais principais da laguna. None

Thermal analysis and microstructural characterization of Mg-Al-Zn system alloys

NASA Astrophysics Data System (ADS)

Król, M.; Tański, T.; Sitek, W.

2015-11-01

The influence of Zn amount and solidification rate on the characteristic temperature of the evaluation of magnesium dendrites during solidification at different cooling rates (0.6-2.5°C) were examined by thermal derivative analysis (TDA). The dendrite coherency point (DCP) is presented with a novel approach based on second derivative cooling curve. Solidification behavior was examined via one thermocouple thermal analysis method. Microstructural assessments were described by optical light microscopy, scanning electron microscopy and energy dispersive X-ray spectroscopy. These studies showed that utilization of d2T/dt2 vs. the time curve methodology provides for analysis of the dendrite coherency point

Rapid enumeration of viable bacteria by image analysis

NASA Technical Reports Server (NTRS)

Singh, A.; Pyle, B. H.; McFeters, G. A.

1989-01-01

A direct viable counting method for enumerating viable bacteria was modified and made compatible with image analysis. A comparison was made between viable cell counts determined by the spread plate method and direct viable counts obtained using epifluorescence microscopy either manually or by automatic image analysis. Cultures of Escherichia coli, Salmonella typhimurium, Vibrio cholerae, Yersinia enterocolitica and Pseudomonas aeruginosa were incubated at 35 degrees C in a dilute nutrient medium containing nalidixic acid. Filtered samples were stained for epifluorescence microscopy and analysed manually as well as by image analysis. Cells enlarged after incubation were considered viable. The viable cell counts determined using image analysis were higher than those obtained by either the direct manual count of viable cells or spread plate methods. The volume of sample filtered or the number of cells in the original sample did not influence the efficiency of the method. However, the optimal concentration of nalidixic acid (2.5-20 micrograms ml-1) and length of incubation (4-8 h) varied with the culture tested. The results of this study showed that under optimal conditions, the modification of the direct viable count method in combination with image analysis microscopy provided an efficient and quantitative technique for counting viable bacteria in a short time.

Analysis of incomplete excisions of basal-cell carcinomas after breadloaf microscopy compared with 3D-microscopy: a prospective randomized and blinded study.

PubMed

Boehringer, Alexandra; Adam, Patrick; Schnabl, Saskia; Häfner, Hans-Martin; Breuninger, Helmut

2015-08-01

Basal-cell carcinomas may show irregular, asymmetric subclinical growth. This study analyzed the efficacy of 'breadloaf' microscopy (serial sectioning) and three-dimensional (3D) microscopy in detecting positive tumor margins. Two hundred eighty-three (283) tumors (51.2%) were put into the breadloaf microscopy group; 270 tumors (48.8%) into the 3D microscopy group. The position of any detected tumor outgrowths was identified in clock face fashion. The time required for cutting and embedding the specimens and the examination of the microscopic slides was measured. Patient/tumor characteristics and surgical margins did not differ significantly. Tumor outgrowths at the excision margin were found in 62 of 283 cases (21.9%) in the breadloaf microscopy group and in 115 of 270 cases (42.6%) in the 3D microscopy group, constituting a highly significant difference (p < 0.001). This difference held true with incomplete excision of fibrosing (infiltrative/sclerosing/morpheaform) tumors [32.9% in the breadloaf microscopy group and 57.5% in the 3D microscopy group (p = 0.003)] and also with solid (nodular) tumors [16.1 and 34.2%, respectively (p < 0.001)]. The mean overall examination time required showed no important difference. In summary, for detection of tumor outgrowths, 3D microscopy has almost twice the sensitivity of breadloaf microscopy, particularly in the situation of aggressive/infiltrative carcinomas. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Confocal laser scanning microscopic photoconversion: a new method to stabilize fluorescently labeled cellular elements for electron microscopic analysis.

PubMed

Colello, Raymond J; Tozer, Jordan; Henderson, Scott C

2012-01-01

Photoconversion, the method by which a fluorescent dye is transformed into a stable, osmiophilic product that can be visualized by electron microscopy, is the most widely used method to enable the ultrastructural analysis of fluorescently labeled cellular structures. Nevertheless, the conventional method of photoconversion using widefield fluorescence microscopy requires long reaction times and results in low-resolution cell targeting. Accordingly, we have developed a photoconversion method that ameliorates these limitations by adapting confocal laser scanning microscopy to the procedure. We have found that this method greatly reduces photoconversion times, as compared to conventional wide field microscopy. Moreover, region-of-interest scanning capabilities of a confocal microscope facilitate the targeting of the photoconversion process to individual cellular or subcellular elements within a fluorescent field. This reduces the area of the cell exposed to light energy, thereby reducing the ultrastructural damage common to this process when widefield microscopes are employed. © 2012 by John Wiley & Sons, Inc.

Super-resolution binding activated localization microscopy through reversible change of DNA conformation.

PubMed

Szczurek, Aleksander; Birk, Udo; Knecht, Hans; Dobrucki, Jurek; Mai, Sabine; Cremer, Christoph

2018-01-01

Methods of super-resolving light microscopy (SRM) have found an exponentially growing range of applications in cell biology, including nuclear structure analyses. Recent developments have proven that Single Molecule Localization Microscopy (SMLM), a type of SRM, is particularly useful for enhanced spatial analysis of the cell nucleus due to its highest resolving capability combined with very specific fluorescent labeling. In this commentary we offer a brief review of the latest methodological development in the field of SMLM of chromatin designated DNA Structure Fluctuation Assisted Binding Activated Localization Microscopy (abbreviated as fBALM) as well as its potential future applications in biology and medicine.

Super-resolution binding activated localization microscopy through reversible change of DNA conformation

PubMed Central

Knecht, Hans; Dobrucki, Jurek; Mai, Sabine

2018-01-01

ABSTRACT Methods of super-resolving light microscopy (SRM) have found an exponentially growing range of applications in cell biology, including nuclear structure analyses. Recent developments have proven that Single Molecule Localization Microscopy (SMLM), a type of SRM, is particularly useful for enhanced spatial analysis of the cell nucleus due to its highest resolving capability combined with very specific fluorescent labeling. In this commentary we offer a brief review of the latest methodological development in the field of SMLM of chromatin designated DNA Structure Fluctuation Assisted Binding Activated Localization Microscopy (abbreviated as fBALM) as well as its potential future applications in biology and medicine. PMID:29297245

Investigation of podosome ring protein arrangement using localization microscopy images.

PubMed

Staszowska, Adela D; Fox-Roberts, Patrick; Foxall, Elizabeth; Jones, Gareth E; Cox, Susan

2017-02-15

Podosomes are adhesive structures formed on the plasma membrane abutting the extracellular matrix of macrophages, osteoclasts, and dendritic cells. They consist of an f-actin core and a ring structure composed of integrins and integrin-associated proteins. The podosome ring plays a major role in adhesion to the underlying extracellular matrix, but its detailed structure is poorly understood. Recently, it has become possible to study the nano-scale structure of podosome rings using localization microscopy. Unlike traditional microscopy images, localization microscopy images are reconstructed using discrete points, meaning that standard image analysis methods cannot be applied. Here, we present a pipeline for podosome identification, protein position calculation, and creating a podosome ring model for use with localization microscopy data. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

A Study of the Effects of High Power Pulsed 2450 MHz Microwaves, ELF modulated Microwaves, and ELF Fields on Human Lymphocytes and Selected Cell Lines

DTIC Science & Technology

1993-01-27

Considerable effect was expended in investigating shifts in intercellular calcium of one particular cell line, Jurket, using flow cytometry methods. No...culture. The following analysis were used to characterize the immortalized cell lines: flow cytometry , electron microscopy, two-dimensional protein gel...further characterized by flow cytometry , electron microscopy, two dimensional protein electrophoresis and nuclear run-off assay. Flow cytometric analysis of

Nano Electronics on Atomically Controlled van der Waals Quantum Heterostructures

DTIC Science & Technology

2015-03-30

for the structural of the atomically sharp interface between hBN and Bi2Te3. Finally, we have developed unprecedentedly clean graphene supercoductor...crystals by MBE method. We also use transmission electron microscopy (TEM) analysis for the structural of the atomically sharp interface between hBN and...by MBE method. We also use transmission electron microscopy (TEM) analysis for the structural of the atomically sharp interface between hBN and Bi2Te3

Atomic force microscopy and scanning electron microscopy evaluation of efficacy of scaling and root planing using magnification: A randomized controlled clinical study

PubMed Central

Mohan, Ranjana; Agrawal, Sudhanshu; Gundappa, Mohan

2013-01-01

Aim: A randomized controlled clinical study was undertaken to evaluate the effectiveness of scaling and root planing (SRP) by using Magnifying Loupes (ML) and dental operating microscope (DOM). Materials and Methods: A total of 90 human teeth scheduled for extraction from 18 patients aged between 25 and 65 years suffering from generalized chronic severe periodontitis were randomly assigned to three treatment groups. Group 1 consisted SRP performed without using magnification (unaided), Group 2-SRP with ML and Group 3-SRP with DOM. Following extractions, samples were prepared for (i) evaluation of surface topography by atomic force microscopy, (ii) presence of smear layer, debris by scanning electron microscopy (iii) elemental analysis by energy dispersive X-ray analysis. Data was subjected to statistical analysis using analysis of variance, post-hoc (Tukey-HSD) and Chi-square test. Results: Statistically significant (P < 0.001) difference was found among the different treatment groups. Group 3 was the best while Group 1 was the least effective technique for SRP. Order of efficacy in terms of the surface was found to be - Palatal < Lingual < Distal ≃ Mesial < Buccal. Efficiency in mandibular to maxillary teeth was found to be significant (P < 0.05), also anterior to posterior teeth (P < 0.05). Conclusion: Magnification tools significantly enhance the efficacy of supragingival and subgingival SRP. PMID:24124292

Experiências internacionais da aplicação de sistemas de apoio à decisão clínica em gastroenterologia

PubMed Central

Tenório, Josceli Maria; Hummel, Anderson Diniz; Sdepanian, Vera Lucia; Pisa, Ivan Torres; de Fátima Marin, Heimar

2015-01-01

Objetivo Descrever as experiências recentes com a aplicação de sistemas de apoio à decisão clínica em gastroenterologia, de forma a estabelecer o nível de desenvolvimento, testes e vantagens conferidas à prática médica com a introdução desses softwares. Métodos Foi realizada busca nas bases de dados PubMed, LILACS e ISI Web of Knowledge, utilizando termos relacionados à sistemas de apoio à decisão e à gastroenterogia, incluindo artigos originais publicados no período entre 2005 e 2010. Foram recuperadas 104 publicações, na busca inicial e, após a aplicação dos critérios de inclusão e exclusão, foram eleitos nove estudos para leitura do texto completo. Resultados Os sistemas de apoio à decisão clínica apresentam grande multiplicidade de problemas clínicos e investigação de doenças. Em 89% dos casos, são descritos modelos experimentais para o desenvolvimento de sistemas de apoio à decisão clínica. A descrição dos resultados obtidos por técnicas de inteligência artificial em 78% das publicações. Em dois dos estudos foram realizadas comparações com o médico e em apenas uma publicação um estudo controlado foi descrito, mostrando evidências de melhorias na prática médica. Conclusão Os estudos mostram potenciais benefícios dos sistemas de apoio à decisão clínica à prática médica, porém, estudos controlados em ambiente real devem ser realizados para comprovar esta perspectiva. PMID:26491625

Elderly with knee osteoarthritis should perform nutritional assessment: integrative literature review.

PubMed

Souza, Isabelle Ferreira da Silva; Oliveira Neta, Rosa Sá de; Gazzola, Juliana Maria; Souza, Marcelo Cardoso de

2017-01-01

To review scientific literature to assess nutritional status of elderly patients with osteoarthritis in the last 16 years. This is an integrative literature review that included articles published in national and international journals indexed in PubMed, SciELO and BIREME. We selected 14 articles, and English language was predominant. The year of publication of articles ranged from 2006 to 2016, and most of papers were cross-sectional studies. To gather papers and for posterior evaluate, we used a validated data collection instrument and the included studies were critical analyzed by reading, gathering and analysis of articles. Studies suggested that there is a positive correlation between obesity and knee osteoarthritis. Obesity is one of the most important modifiable factors in worsening of osteoarthritis symptoms. RESUMO O objetivo da pesquisa foi revisar a produção científica referente à avaliação do estado nutricional de idosos com osteoartrite nos últimos 16 anos. Assim, o estudo foi uma revisão integrativa da literatura, realizada com a busca de artigos publicados em periódicos nacionais e internacionais indexados no PubMed, na SciELO e na BIREME. Foram selecionados 14 artigos, e o idioma inglês foi preponderante. O período de publicação dos artigos variou de 2006 a 2016, com predominância de estudos do tipo transversais. Para reunir os artigos e para posterior avaliação, foi utilizado um instrumento de coleta de dados validado, e as análises críticas dos estudos incluídos foram realizadas por meio da leitura, do agrupamento e da análise dos artigos. As pesquisas sugeriram que existe correlação positiva entre obesidade e osteoartrite de joelhos. Além disto, a obesidade é um dos fatores mais significativos e modificáveis no agravamento dos sintomas da osteoartrite.

Asbestos Analysis: What School Officials Should Know.

ERIC Educational Resources Information Center

Harris, Bonnie Lee

1984-01-01

Transmission electron microscopy and scanning electron microscopy are used to detect asbestos by analyzing filters from air tests. The modes of operation and types of samples examined by each are explained. Circumstances that a school board should consider when deciding whether to use these methods are discussed. (MLF)

Scanning electron microscopy analysis of corrosion degradation on tinplate substrates.

PubMed

Zumelzu, E; Cabezas, C; Vera, A

2003-01-01

The degradation of electrolytic tinplate used in food containers was analysed and evaluated, using scanning electron microscopy and electrochemical measurements of microcorrosion and ion dissolution by atomic absorption to prevent food contamination caused by metal traces and to increase the durability of such tinplates.

Confocal Raman Microscopy: new perspective on the weathering of anhydrous cement

NASA Astrophysics Data System (ADS)

Torres-Carrasco, M.; del Campo, A.; de la Rubia, MA; Reyes, E.; Moragues, A.; Fernández, JF

2017-10-01

Raman spectroscopy when is combined with Confocal microscopy is a non-destructive technique that allow us to obtain information in cementitious materials. In this study, we present non-destructive image and structural analysis of anhydrous cement with carbonation evidences by Confocal Raman Microscopy (CRM). The results obtained by CRM show a direct relationship between the presence of the weathering processes of an anhydrous cement with the presence of sulphates and surprisingly, with the existence of amorphous carbon in the medium.

Full information acquisition in scanning probe microscopy and spectroscopy

DOEpatents

Jesse, Stephen; Belianinov, Alex; Kalinin, Sergei V.; Somnath, Suhas

2017-04-04

Apparatus and methods are described for scanning probe microscopy and spectroscopy based on acquisition of full probe response. The full probe response contains valuable information about the probe-sample interaction that is lost in traditional scanning probe microscopy and spectroscopy methods. The full probe response is analyzed post data acquisition using fast Fourier transform and adaptive filtering, as well as multivariate analysis. The full response data is further compressed to retain only statistically significant components before being permanently stored.

Evaluation of mobile digital light-emitting diode fluorescence microscopy in Hanoi, Viet Nam.

PubMed

Chaisson, L H; Reber, C; Phan, H; Switz, N; Nilsson, L M; Myers, F; Nhung, N V; Luu, L; Pham, T; Vu, C; Nguyen, H; Nguyen, A; Dinh, T; Nahid, P; Fletcher, D A; Cattamanchi, A

2015-09-01

Hanoi Lung Hospital, Hanoi, Viet Nam. To compare the accuracy of CellScopeTB, a manually operated mobile digital fluorescence microscope, with conventional microscopy techniques. Patients referred for sputum smear microscopy to the Hanoi Lung Hospital from May to September 2013 were included. Ziehl-Neelsen (ZN) smear microscopy, conventional light-emitting diode (LED) fluorescence microscopy (FM), CellScopeTB-based LED FM and Xpert(®) MTB/RIF were performed on sputum samples. The sensitivity and specificity of microscopy techniques were determined in reference to Xpert results, and differences were compared using McNemar's paired test of proportions. Of 326 patients enrolled, 93 (28.5%) were Xpert-positive for TB. The sensitivity of ZN microscopy, conventional LED FM, and CellScopeTB-based LED FM was respectively 37.6% (95%CI 27.8-48.3), 41.9% (95%CI 31.8-52.6), and 35.5% (95%CI 25.8-46.1). The sensitivity of CellScopeTB was similar to that of conventional LED FM (difference -6.5%, 95%CI -18.2 to 5.3, P = 0.33) and ZN microscopy (difference -2.2%, 95%CI -9.2 to 4.9, P = 0.73). The specificity was >99% for all three techniques. CellScopeTB performed similarly to conventional microscopy techniques in the hands of experienced TB microscopists. However, the sensitivity of all sputum microscopy techniques was low. Options enabled by digital microscopy, such as automated imaging with real-time computerized analysis, should be explored to increase sensitivity.

Quantification of surface displacements and electromechanical phenomena via dynamic atomic force microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Balke, Nina; Jesse, Stephen; Yu, Pu

Detection of dynamic surface displacements associated with local changes in material strain provides access to a number of phenomena and material properties. Contact resonance-enhanced methods of atomic force microscopy (AFM) have been shown capable of detecting ~1–3 pm-level surface displacements, an approach used in techniques such as piezoresponse force microscopy, atomic force acoustic microscopy, and ultrasonic force microscopy. Here, based on an analytical model of AFM cantilever vibrations, we demonstrate a guideline to quantify surface displacements with high accuracy by taking into account the cantilever shape at the first resonant contact mode, depending on the tip–sample contact stiffness. The approachmore » has been experimentally verified and further developed for piezoresponse force microscopy (PFM) using well-defined ferroelectric materials. These results open up a way to accurate and precise measurements of surface displacement as well as piezoelectric constants at the pm-scale with nanometer spatial resolution and will allow avoiding erroneous data interpretations and measurement artifacts. Furthermore, this analysis is directly applicable to all cantilever-resonance-based scanning probe microscopy (SPM) techniques.« less

Quantification of surface displacements and electromechanical phenomena via dynamic atomic force microscopy

DOE PAGES

Balke, Nina; Jesse, Stephen; Yu, Pu; ...

2016-09-15

Detection of dynamic surface displacements associated with local changes in material strain provides access to a number of phenomena and material properties. Contact resonance-enhanced methods of atomic force microscopy (AFM) have been shown capable of detecting ~1–3 pm-level surface displacements, an approach used in techniques such as piezoresponse force microscopy, atomic force acoustic microscopy, and ultrasonic force microscopy. Here, based on an analytical model of AFM cantilever vibrations, we demonstrate a guideline to quantify surface displacements with high accuracy by taking into account the cantilever shape at the first resonant contact mode, depending on the tip–sample contact stiffness. The approachmore » has been experimentally verified and further developed for piezoresponse force microscopy (PFM) using well-defined ferroelectric materials. These results open up a way to accurate and precise measurements of surface displacement as well as piezoelectric constants at the pm-scale with nanometer spatial resolution and will allow avoiding erroneous data interpretations and measurement artifacts. Furthermore, this analysis is directly applicable to all cantilever-resonance-based scanning probe microscopy (SPM) techniques.« less

New approaches in renal microscopy: volumetric imaging and superresolution microscopy.

PubMed

Kim, Alfred H J; Suleiman, Hani; Shaw, Andrey S

2016-05-01

Histologic and electron microscopic analysis of the kidney has provided tremendous insight into structures such as the glomerulus and nephron. Recent advances in imaging, such as deep volumetric approaches and superresolution microscopy, have the capacity to dramatically enhance our current understanding of the structure and function of the kidney. Volumetric imaging can generate images millimeters below the surface of the intact kidney. Superresolution microscopy breaks the diffraction barrier inherent in traditional light microscopy, enabling the visualization of fine structures. Here, we describe new approaches to deep volumetric and superresolution microscopy of the kidney. Rapid advances in lasers, microscopic objectives, and tissue preparation have transformed our ability to deep volumetric image the kidney. Innovations in sample preparation have allowed for superresolution imaging with electron microscopy correlation, providing unprecedented insight into the structures within the glomerulus. Technological advances in imaging have revolutionized our capacity to image both large volumes of tissue and the finest structural details of a cell. These new advances have the potential to provide additional profound observations into the normal and pathologic functions of the kidney.

Working at the microscope: analysis of the activities involved in diagnostic pathology.

PubMed

Randell, Rebecca; Ruddle, Roy A; Quirke, Phil; Thomas, Rhys G; Treanor, Darren

2012-02-01

To study the current work practice of histopathologists to inform the design of digital microscopy systems. Four gastrointestinal histopathologists were video-recorded as they undertook their routine work. Analysis of the video data shows a range of activities beyond viewing slides involved in reporting a case. There is much overlapping of activities, supported by the 'eyes free' nature of the pathologists' interaction with the microscope. The order and timing of activities varies according to consultant. In order to support the work of pathologists adequately, digital microscopy systems need to provide support for a range of activities beyond viewing slides. Digital microscopy systems should support multitasking, while also providing flexibility so that pathologists can adapt their use of the technology to their own working patterns. © 2011 Blackwell Publishing Ltd.

FT Raman microscopy of untreated natural plant fibres

NASA Astrophysics Data System (ADS)

Edwards, H. G. M.; Farwell, D. W.; Webster, D.

1997-11-01

The application of FT-Raman microscopy to the non-destructive analysis of natural plant fibres is demonstrated with samples of flax, jute, ramie, cotton, kapok, sisal and coconut fibre. Vibrational assignments are proposed and characteristic features of each material are presented. Samples were not pre-treated chemically before analysis and were used directly from their respective storage collection; the adaptation of the Raman microscopic technique to the identification of specimens of natural fibres in archaeological burial sites is explored for its forensic potential.

Synthesis of samarium doped gadolinium oxide nanorods, its spectroscopic and physical properties

NASA Astrophysics Data System (ADS)

Boopathi, G.; Gokul Raj, S.; Ramesh Kumar, G.; Mohan, R.; Mohan, S.

2018-06-01

One-dimensional samarium doped gadolinium oxide [Sm:Gd2O3] nanorods have been synthesized successfully through co-precipitation technique in aqueous solution. The as-synthesized and calcined products were characterized by using powder X-ray diffraction pattern, Fourier transform Raman spectroscopy, thermogravimetric/differential thermal analysis, scanning electron microscopy with energy-dispersive X-ray analysis, transmission electron microscopy, Fourier transform infrared spectroscopy, Ultraviolet-Visible spectrometry, photoluminescence spectrophotometer and X-ray photoelectron spectroscopy techniques. The obtained results are discussed in detailed manner.

Point-of-care mobile digital microscopy and deep learning for the detection of soil-transmitted helminths and Schistosoma haematobium.

PubMed

Holmström, Oscar; Linder, Nina; Ngasala, Billy; Mårtensson, Andreas; Linder, Ewert; Lundin, Mikael; Moilanen, Hannu; Suutala, Antti; Diwan, Vinod; Lundin, Johan

2017-06-01

Microscopy remains the gold standard in the diagnosis of neglected tropical diseases. As resource limited, rural areas often lack laboratory equipment and trained personnel, new diagnostic techniques are needed. Low-cost, point-of-care imaging devices show potential in the diagnosis of these diseases. Novel, digital image analysis algorithms can be utilized to automate sample analysis. Evaluation of the imaging performance of a miniature digital microscopy scanner for the diagnosis of soil-transmitted helminths and Schistosoma haematobium, and training of a deep learning-based image analysis algorithm for automated detection of soil-transmitted helminths in the captured images. A total of 13 iodine-stained stool samples containing Ascaris lumbricoides, Trichuris trichiura and hookworm eggs and 4 urine samples containing Schistosoma haematobium were digitized using a reference whole slide-scanner and the mobile microscopy scanner. Parasites in the images were identified by visual examination and by analysis with a deep learning-based image analysis algorithm in the stool samples. Results were compared between the digital and visual analysis of the images showing helminth eggs. Parasite identification by visual analysis of digital slides captured with the mobile microscope was feasible for all analyzed parasites. Although the spatial resolution of the reference slide-scanner is higher, the resolution of the mobile microscope is sufficient for reliable identification and classification of all parasites studied. Digital image analysis of stool sample images captured with the mobile microscope showed high sensitivity for detection of all helminths studied (range of sensitivity = 83.3-100%) in the test set (n = 217) of manually labeled helminth eggs. In this proof-of-concept study, the imaging performance of a mobile, digital microscope was sufficient for visual detection of soil-transmitted helminths and Schistosoma haematobium. Furthermore, we show that deep learning-based image analysis can be utilized for the automated detection and classification of helminths in the captured images.

Point-of-care mobile digital microscopy and deep learning for the detection of soil-transmitted helminths and Schistosoma haematobium

PubMed Central

Holmström, Oscar; Linder, Nina; Ngasala, Billy; Mårtensson, Andreas; Linder, Ewert; Lundin, Mikael; Moilanen, Hannu; Suutala, Antti; Diwan, Vinod; Lundin, Johan

2017-01-01

ABSTRACT Background: Microscopy remains the gold standard in the diagnosis of neglected tropical diseases. As resource limited, rural areas often lack laboratory equipment and trained personnel, new diagnostic techniques are needed. Low-cost, point-of-care imaging devices show potential in the diagnosis of these diseases. Novel, digital image analysis algorithms can be utilized to automate sample analysis. Objective: Evaluation of the imaging performance of a miniature digital microscopy scanner for the diagnosis of soil-transmitted helminths and Schistosoma haematobium, and training of a deep learning-based image analysis algorithm for automated detection of soil-transmitted helminths in the captured images. Methods: A total of 13 iodine-stained stool samples containing Ascaris lumbricoides, Trichuris trichiura and hookworm eggs and 4 urine samples containing Schistosoma haematobium were digitized using a reference whole slide-scanner and the mobile microscopy scanner. Parasites in the images were identified by visual examination and by analysis with a deep learning-based image analysis algorithm in the stool samples. Results were compared between the digital and visual analysis of the images showing helminth eggs. Results: Parasite identification by visual analysis of digital slides captured with the mobile microscope was feasible for all analyzed parasites. Although the spatial resolution of the reference slide-scanner is higher, the resolution of the mobile microscope is sufficient for reliable identification and classification of all parasites studied. Digital image analysis of stool sample images captured with the mobile microscope showed high sensitivity for detection of all helminths studied (range of sensitivity = 83.3–100%) in the test set (n = 217) of manually labeled helminth eggs. Conclusions: In this proof-of-concept study, the imaging performance of a mobile, digital microscope was sufficient for visual detection of soil-transmitted helminths and Schistosoma haematobium. Furthermore, we show that deep learning-based image analysis can be utilized for the automated detection and classification of helminths in the captured images. PMID:28838305

Analysis of human knee osteoarthritic cartilage using polarization sensitive second harmonic generation microscopy

NASA Astrophysics Data System (ADS)

Kumar, Rajesh; Grønhaug, Kirsten M.; Romijn, Elisabeth I.; Drogset, Jon O.; Lilledahl, Magnus B.

2014-05-01

Osteoarthritis is one of the most prevalent joint diseases in the world. Although the cause of osteoarthritis is not exactly clear, the disease results in a degradation of the quality of the articular cartilage including collagen and other extracellular matrix components. We have investigated alterations in the structure of collagen fibers in the cartilage tissue of the human knee using mulitphoton microscopy. Due to inherent high nonlinear susceptibility, ordered collagen fibers present in the cartilage tissue matrix produces strong second harmonic generation (SHG) signals. Significant morphological differences are found in different Osteoarthritic grades of cartilage by SHG microscopy. Based on the polarization analysis of the SHG signal, we find that a few locations of hyaline cartilage (mainly type II collagen) is being replaced by fibrocartilage (mainly type I cartilage), in agreement with earlier literature. To locate the different types and quantify the alteration in the structure of collagen fiber, we employ polarization-SHG microscopic analysis, also referred to as _-tensor imaging. The image analysis of p-SHG image obtained by excitation polarization measurements would represent different tissue constituents with different numerical values at pixel level resolution.

Web-based visualisation and analysis of 3D electron-microscopy data from EMDB and PDB.

PubMed

Lagerstedt, Ingvar; Moore, William J; Patwardhan, Ardan; Sanz-García, Eduardo; Best, Christoph; Swedlow, Jason R; Kleywegt, Gerard J

2013-11-01

The Protein Data Bank in Europe (PDBe) has developed web-based tools for the visualisation and analysis of 3D electron microscopy (3DEM) structures in the Electron Microscopy Data Bank (EMDB) and Protein Data Bank (PDB). The tools include: (1) a volume viewer for 3D visualisation of maps, tomograms and models, (2) a slice viewer for inspecting 2D slices of tomographic reconstructions, and (3) visual analysis pages to facilitate analysis and validation of maps, tomograms and models. These tools were designed to help non-experts and experts alike to get some insight into the content and assess the quality of 3DEM structures in EMDB and PDB without the need to install specialised software or to download large amounts of data from these archives. The technical challenges encountered in developing these tools, as well as the more general considerations when making archived data available to the user community through a web interface, are discussed. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Invited Review Article: Imaging techniques for harmonic and multiphoton absorption fluorescence microscopy

PubMed Central

Carriles, Ramón; Schafer, Dawn N.; Sheetz, Kraig E.; Field, Jeffrey J.; Cisek, Richard; Barzda, Virginijus; Sylvester, Anne W.; Squier, Jeffrey A.

2009-01-01

We review the current state of multiphoton microscopy. In particular, the requirements and limitations associated with high-speed multiphoton imaging are considered. A description of the different scanning technologies such as line scan, multifoci approaches, multidepth microscopy, and novel detection techniques is given. The main nonlinear optical contrast mechanisms employed in microscopy are reviewed, namely, multiphoton excitation fluorescence, second harmonic generation, and third harmonic generation. Techniques for optimizing these nonlinear mechanisms through a careful measurement of the spatial and temporal characteristics of the focal volume are discussed, and a brief summary of photobleaching effects is provided. Finally, we consider three new applications of multiphoton microscopy: nonlinear imaging in microfluidics as applied to chemical analysis and the use of two-photon absorption and self-phase modulation as contrast mechanisms applied to imaging problems in the medical sciences. PMID:19725639

Tendências De Teses e Dissertações Sobre Educação em Astronomia No Brasil

NASA Astrophysics Data System (ADS)

Bretones, Paulo Sergio; Megid Neto, Jorge

2005-07-01

Apresentam-se os resultados de uma pesquisa do tipo estado da arte sobre teses e dissertações defendidas no Brasil e relativas ao ensino de Astronomia, com objetivo de identificar essa produção e conhecer as principais tendências da pesquisa nesse campo. Foram localizadas 13 dissertações de mestrado e 3 teses de doutorado, as quais foram estudadas em função dos seguintes aspectos: isntituição, ano de defesa, nível escolar abrangido no estudo, foco temático do estudo e gênero de trabalho acadêmico. Pretende-se assim colaborar com a divulgação ampla da produção acadêmica na área. Ao mesmo tempo o estudo possibilita, a partir de investigações decorrentes, apontar as contribuições dessa produção para o ensino e sinalizar com necessidades a serem supridas por futuras pesquisas.

A fast global fitting algorithm for fluorescence lifetime imaging microscopy based on image segmentation.

PubMed

Pelet, S; Previte, M J R; Laiho, L H; So, P T C

2004-10-01

Global fitting algorithms have been shown to improve effectively the accuracy and precision of the analysis of fluorescence lifetime imaging microscopy data. Global analysis performs better than unconstrained data fitting when prior information exists, such as the spatial invariance of the lifetimes of individual fluorescent species. The highly coupled nature of global analysis often results in a significantly slower convergence of the data fitting algorithm as compared with unconstrained analysis. Convergence speed can be greatly accelerated by providing appropriate initial guesses. Realizing that the image morphology often correlates with fluorophore distribution, a global fitting algorithm has been developed to assign initial guesses throughout an image based on a segmentation analysis. This algorithm was tested on both simulated data sets and time-domain lifetime measurements. We have successfully measured fluorophore distribution in fibroblasts stained with Hoechst and calcein. This method further allows second harmonic generation from collagen and elastin autofluorescence to be differentiated in fluorescence lifetime imaging microscopy images of ex vivo human skin. On our experimental measurement, this algorithm increased convergence speed by over two orders of magnitude and achieved significantly better fits. Copyright 2004 Biophysical Society

Open-source do-it-yourself multi-color fluorescence smartphone microscopy

PubMed Central

Sung, Yulung; Campa, Fernando; Shih, Wei-Chuan

2017-01-01

Fluorescence microscopy is an important technique for cellular and microbiological investigations. Translating this technique onto a smartphone can enable particularly powerful applications such as on-site analysis, on-demand monitoring, and point-of-care diagnostics. Current fluorescence smartphone microscope setups require precise illumination and imaging alignment which altogether limit its broad adoption. We report a multi-color fluorescence smartphone microscope with a single contact lens-like add-on lens and slide-launched total-internal-reflection guided illumination for three common tasks in investigative fluorescence microscopy: autofluorescence, fluorescent stains, and immunofluorescence. The open-source, simple and cost-effective design has the potential for do-it-yourself fluorescence smartphone microscopy. PMID:29188104

Optofluidic time-stretch microscopy: recent advances

NASA Astrophysics Data System (ADS)

Lei, Cheng; Nitta, Nao; Ozeki, Yasuyuki; Goda, Keisuke

2018-06-01

Flow cytometry is an indispensable method for valuable applications in numerous fields such as immunology, pathology, pharmacology, molecular biology, and marine biology. Optofluidic time-stretch microscopy is superior to conventional flow cytometry methods for its capability to acquire high-quality images of single cells at a high-throughput exceeding 10,000 cells per second. This makes it possible to extract copious information from cellular images for accurate cell detection and analysis with the assistance of machine learning. Optofluidic time-stretch microscopy has proven its effectivity in various applications, including microalga-based biofuel production, evaluation of thrombotic disorders, as well as drug screening and discovery. In this review, we discuss the principles and recent advances of optofluidic time-stretch microscopy.

Frequency domain phase-shifted confocal microscopy (FDPCM) with array detection

NASA Astrophysics Data System (ADS)

Ge, Baoliang; Huang, Yujia; Fang, Yue; Kuang, Cuifang; Xiu, Peng; Liu, Xu

2017-09-01

We proposed a novel method to reconstruct images taken by array detected confocal microscopy without prior knowledge about its detector distribution. The proposed frequency domain phase-shifted confocal microscopy (FDPCM) shifts the image from each detection channel to its corresponding place by substituting the phase information in Fourier domain. Theoretical analysis shows that our method could approach the resolution nearly twofold of wide-field microscopy. Simulation and experiment results are also shown to verify the applicability and effectiveness of our method. Compared to Airyscan, our method holds the advantage of simplicity and convenience to be applied to array detectors with different structure, which makes FDPCM have great potential in the application of biomedical observation in the future.

Optofluidic time-stretch microscopy: recent advances

NASA Astrophysics Data System (ADS)

Lei, Cheng; Nitta, Nao; Ozeki, Yasuyuki; Goda, Keisuke

2018-04-01

Flow cytometry is an indispensable method for valuable applications in numerous fields such as immunology, pathology, pharmacology, molecular biology, and marine biology. Optofluidic time-stretch microscopy is superior to conventional flow cytometry methods for its capability to acquire high-quality images of single cells at a high-throughput exceeding 10,000 cells per second. This makes it possible to extract copious information from cellular images for accurate cell detection and analysis with the assistance of machine learning. Optofluidic time-stretch microscopy has proven its effectivity in various applications, including microalga-based biofuel production, evaluation of thrombotic disorders, as well as drug screening and discovery. In this review, we discuss the principles and recent advances of optofluidic time-stretch microscopy.

Airborne asbestos in Colorado public schools.

PubMed

Chadwick, D A; Buchan, R M; Beaulieu, H J

1985-02-01

Levels of airborne asbestos for six Colorado public school facilities with sprayed-on asbestos materials were documented using three analytical techniques. Phase contrast microscopy showed levels up to the thousandths of a fiber per cubic centimeter (f/cc), scanning electron microscopy (SEM) up to the hundredths of a f/cc, and transmission electron microscopy coupled to selected area electron diffraction and energy dispersive X-ray analysis (TEM-SAED-EDXA) up to the tenths of an asbestos f/cc. Phase contrast microscopy was found to be an inadequate analytical technique for documenting the levels of airborne asbestos fibers in the schools: only large fibers which were not embedded in the filter were counted, and asbestos fibers were not distinguished from nonasbestos.

Imaging chromatin nanostructure with binding-activated localization microscopy based on DNA structure fluctuations

PubMed Central

Szczurek, Aleksander; Klewes, Ludger; Xing, Jun; Gourram, Amine; Birk, Udo; Knecht, Hans; Dobrucki, Jurek W.; Mai, Sabine

2017-01-01

Abstract Advanced light microscopy is an important tool for nanostructure analysis of chromatin. In this report we present a general concept for Single Molecule localization Microscopy (SMLM) super-resolved imaging of DNA-binding dyes based on modifying the properties of DNA and the dye. By careful adjustment of the chemical environment leading to local, reversible DNA melting and hybridization control over the fluorescence signal of the DNA-binding dye molecules can be introduced. We postulate a transient binding as the basis for our variation of binding-activated localization microscopy (BALM). We demonstrate that several intercalating and minor-groove binding DNA dyes can be used to register (optically isolate) only a few DNA-binding dye signals at a time. To highlight this DNA structure fluctuation-assisted BALM (fBALM), we applied it to measure, for the first time, nanoscale differences in nuclear architecture in model ischemia with an anticipated structural resolution of approximately 50 nm. Our data suggest that this approach may open an avenue for the enhanced microscopic analysis of chromatin nano-architecture and hence the microscopic analysis of nuclear structure aberrations occurring in various pathological conditions. It may also become possible to analyse nuclear nanostructure differences in different cell types, stages of development or environmental stress conditions. PMID:28082388

Correlative scanning-transmission electron microscopy reveals that a chimeric flavivirus is released as individual particles in secretory vesicles.

PubMed

Burlaud-Gaillard, Julien; Sellin, Caroline; Georgeault, Sonia; Uzbekov, Rustem; Lebos, Claude; Guillaume, Jean-Marc; Roingeard, Philippe

2014-01-01

The intracellular morphogenesis of flaviviruses has been well described, but flavivirus release from the host cell remains poorly documented. We took advantage of the optimized production of an attenuated chimeric yellow fever/dengue virus for vaccine purposes to study this phenomenon by microscopic approaches. Scanning electron microscopy (SEM) showed the release of numerous viral particles at the cell surface through a short-lived process. For transmission electron microscopy (TEM) studies of the intracellular ultrastructure of the small number of cells releasing viral particles at a given time, we developed a new correlative microscopy method: CSEMTEM (for correlative scanning electron microscopy - transmission electron microscopy). CSEMTEM analysis suggested that chimeric flavivirus particles were released as individual particles, in small exocytosis vesicles, via a regulated secretory pathway. Our morphological findings provide new insight into interactions between flaviviruses and cells and demonstrate that CSEMTEM is a useful new method, complementary to SEM observations of biological events by intracellular TEM investigations.

Correlative Scanning-Transmission Electron Microscopy Reveals that a Chimeric Flavivirus Is Released as Individual Particles in Secretory Vesicles

PubMed Central

Burlaud-Gaillard, Julien; Sellin, Caroline; Georgeault, Sonia; Uzbekov, Rustem; Lebos, Claude; Guillaume, Jean-Marc; Roingeard, Philippe

2014-01-01

The intracellular morphogenesis of flaviviruses has been well described, but flavivirus release from the host cell remains poorly documented. We took advantage of the optimized production of an attenuated chimeric yellow fever/dengue virus for vaccine purposes to study this phenomenon by microscopic approaches. Scanning electron microscopy (SEM) showed the release of numerous viral particles at the cell surface through a short-lived process. For transmission electron microscopy (TEM) studies of the intracellular ultrastructure of the small number of cells releasing viral particles at a given time, we developed a new correlative microscopy method: CSEMTEM (for correlative scanning electron microscopy - transmission electron microscopy). CSEMTEM analysis suggested that chimeric flavivirus particles were released as individual particles, in small exocytosis vesicles, via a regulated secretory pathway. Our morphological findings provide new insight into interactions between flaviviruses and cells and demonstrate that CSEMTEM is a useful new method, complementary to SEM observations of biological events by intracellular TEM investigations. PMID:24681578

Boundary segmentation for fluorescence microscopy using steerable filters

NASA Astrophysics Data System (ADS)

Ho, David Joon; Salama, Paul; Dunn, Kenneth W.; Delp, Edward J.

2017-02-01

Fluorescence microscopy is used to image multiple subcellular structures in living cells which are not readily observed using conventional optical microscopy. Moreover, two-photon microscopy is widely used to image structures deeper in tissue. Recent advancement in fluorescence microscopy has enabled the generation of large data sets of images at different depths, times, and spectral channels. Thus, automatic object segmentation is necessary since manual segmentation would be inefficient and biased. However, automatic segmentation is still a challenging problem as regions of interest may not have well defined boundaries as well as non-uniform pixel intensities. This paper describes a method for segmenting tubular structures in fluorescence microscopy images of rat kidney and liver samples using adaptive histogram equalization, foreground/background segmentation, steerable filters to capture directional tendencies, and connected-component analysis. The results from several data sets demonstrate that our method can segment tubular boundaries successfully. Moreover, our method has better performance when compared to other popular image segmentation methods when using ground truth data obtained via manual segmentation.

Use of the polymerase chain reaction to directly detect malaria parasites in blood samples from the Venezuelan Amazon.

PubMed

Laserson, K F; Petralanda, I; Hamlin, D M; Almera, R; Fuentes, M; Carrasquel, A; Barker, R H

1994-02-01

We have examined the reproducibility, sensitivity, and specificity of detecting Plasmodium falciparum using the polymerase chain reaction (PCR) and the species-specific probe pPF14 under field conditions in the Venezuelan Amazon. Up to eight samples were field collected from each of 48 consenting Amerindians presenting with symptoms of malaria. Sample processing and analysis was performed at the Centro Amazonico para la Investigacion y Control de Enfermedades Tropicales Simon Bolivar. A total of 229 samples from 48 patients were analyzed by PCR methods using four different P. falciparum-specific probes. One P. vivax-specific probe and by conventional microscopy. Samples in which results from PCR and microscopy differed were reanalyzed at a higher sensitivity by microscopy. Results suggest that microscopy-negative, PCR-positive samples are true positives, and that microscopy-positive and PCR-negative samples are true negatives. The sensitivity of the DNA probe/PCR method was 78% and its specificity was 97%. The positive predictive value of the PCR method was 88%, and the negative predictive value was 95%. Through the analysis of multiple blood samples from each individual, the DNA probe/PCR methodology was found to have an inherent reproducibility that was highly statistically significant.

Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells.

PubMed

Hofemeier, Arne D; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F W; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

2016-05-26

Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO4(3-) symmetric stretch vibrations at 959 cm(-1) assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue-implant-interfaces or disease diagnosis.

Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells

NASA Astrophysics Data System (ADS)

Hofemeier, Arne D.; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F. W.; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

2016-05-01

Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO43- symmetric stretch vibrations at 959 cm-1 assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue-implant-interfaces or disease diagnosis.

Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells

PubMed Central

Hofemeier, Arne D.; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F. W.; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

2016-01-01

Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO43− symmetric stretch vibrations at 959 cm−1 assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue–implant-interfaces or disease diagnosis. PMID:27225821

IMIS: An intelligence microscope imaging system

NASA Technical Reports Server (NTRS)

Caputo, Michael; Hunter, Norwood; Taylor, Gerald

1994-01-01

Until recently microscope users in space relied on traditional microscopy techniques that required manual operation of the microscope and recording of observations in the form of written notes, drawings, or photographs. This method was time consuming and required the return of film and drawings from space for analysis. No real-time data analysis was possible. Advances in digital and video technologies along with recent developments in article intelligence will allow future space microscopists to have a choice of three additional modes of microscopy: remote coaching, remote control, and automation. Remote coaching requires manual operations of the microscope with instructions given by two-way audio/video transmission during critical phases of the experiment. When using the remote mode of microscopy, the Principal Investigator controls the microscope from the ground. The automated mode employs artificial intelligence to control microscope functions and is the only mode that can be operated in the other three modes as well. The purpose of this presentation is to discuss the advantages and disadvantages of the four modes of of microscopy and how the IMIS, a proposed intelligent microscope imaging system, can be used as a model for developing and testing concepts, operating procedures, and equipment design of specifications required to provide a comprehensive microscopy/imaging capability onboard Space Station Freedom.

Full-field optical coherence microscopy is a novel technique for imaging enteric ganglia in the gastrointestinal tract

PubMed Central

CORON, E.; AUKSORIUS, E.; PIERETTI, A.; MAHÉ, M. M.; LIU, L.; STEIGER, C.; BROMBERG, Y.; BOUMA, B.; TEARNEY, G.; NEUNLIST, M.; GOLDSTEIN, A. M.

2013-01-01

Background Noninvasive methods are needed to improve the diagnosis of enteric neuropathies. Full-field optical coherence microscopy (FFOCM) is a novel optical microscopy modality that can acquire 1 μm resolution images of tissue. The objective of this research was to demonstrate FFOCM imaging for the characterization of the enteric nervous system (ENS). Methods Normal mice and EdnrB−/− mice, a model of Hirschsprung’s disease (HD), were imaged in three-dimensions ex vivo using FFOCM through the entire thickness and length of the gut. Quantitative analysis of myenteric ganglia was performed on FFOCM images obtained from whole-mount tissues and compared with immunohistochemistry imaged by confocal microscopy. Key Results Full-field optical coherence microscopy enabled visualization of the full thickness gut wall from serosa to mucosa. Images of the myenteric plexus were successfully acquired from the stomach, duodenum, colon, and rectum. Quantification of ganglionic neuronal counts on FFOCM images revealed strong interobserver agreement and identical values to those obtained by immunofluorescence microscopy. In EdnrB−/− mice, FFOCM analysis revealed a significant decrease in ganglia density along the colorectum and a significantly lower density of ganglia in all colorectal segments compared with normal mice. Conclusions & Inferences Full-field optical coherence microscopy enables optical microscopic imaging of the ENS within the bowel wall along the entire intestine. FFOCM is able to differentiate ganglionic from aganglionic colon in a mouse model of HD, and can provide quantitative assessment of ganglionic density. With further refinements that enable bowel wall imaging in vivo, this technology has the potential to revolutionize the characterization of the ENS and the diagnosis of enteric neuropathies. PMID:23106847

Applications of microscopy in Salmonella research.

PubMed

Malt, Layla M; Perrett, Charlotte A; Humphrey, Suzanne; Jepson, Mark A

2015-01-01

Salmonella enterica is a Gram-negative enteropathogen that can cause localized infections, typically resulting in gastroenteritis, or systemic infection, e.g., typhoid fever, in humans and many other animals. Understanding the mechanisms by which Salmonella induces disease has been the focus of intensive research. This has revealed that Salmonella invasion requires dynamic cross-talk between the microbe and host cells, in which bacterial adherence rapidly leads to a complex sequence of cellular responses initiated by proteins translocated into the host cell by a type 3 secretion system. Once these Salmonella-induced responses have resulted in bacterial invasion, proteins translocated by a second type 3 secretion system initiate further modulation of cellular activities to enable survival and replication of the invading pathogen. Elucidation of the complex and highly dynamic pathogen-host interactions ultimately requires analysis at the level of single cells and single infection events. To achieve this goal, researchers have applied a diverse range of microscopy techniques to analyze Salmonella infection in models ranging from whole animal to isolated cells and simple eukaryotic organisms. For example, electron microscopy and high-resolution light microscopy techniques such as confocal microscopy can reveal the precise location of Salmonella and its relationship to cellular components. Widefield light microscopy is a simpler approach with which to study the interaction of bacteria with host cells and often has advantages for live cell imaging, enabling detailed analysis of the dynamics of infection and cellular responses. Here we review the use of imaging techniques in Salmonella research and compare the capabilities of different classes of microscope to address specific types of research question. We also provide protocols and notes on some microscopy techniques used routinely in our own research.

Analysis of leaf surfaces using scanning ion conductance microscopy.

PubMed

Walker, Shaun C; Allen, Stephanie; Bell, Gordon; Roberts, Clive J

2015-05-01

Leaf surfaces are highly complex functional systems with well defined chemistry and structure dictating the barrier and transport properties of the leaf cuticle. It is a significant imaging challenge to analyse the very thin and often complex wax-like leaf cuticle morphology in their natural state. Scanning electron microscopy (SEM) and to a lesser extent Atomic force microscopy are techniques that have been used to study the leaf surface but their remains information that is difficult to obtain via these approaches. SEM is able to produce highly detailed and high-resolution images needed to study leaf structures at the submicron level. It typically operates in a vacuum or low pressure environment and as a consequence is generally unable to deal with the in situ analysis of dynamic surface events at submicron scales. Atomic force microscopy also possess the high-resolution imaging required and can follow dynamic events in ambient and liquid environments, but can over exaggerate small features and cannot image most leaf surfaces due to their inherent roughness at the micron scale. Scanning ion conductance microscopy (SICM), which operates in a liquid environment, provides a potential complementary analytical approach able to address these issues and which is yet to be explored for studying leaf surfaces. Here we illustrate the potential of SICM on various leaf surfaces and compare the data to SEM and atomic force microscopy images on the same samples. In achieving successful imaging we also show that SICM can be used to study the wetting of hydrophobic surfaces in situ. This has potentially wider implications than the study of leaves alone as surface wetting phenomena are important in a range of fundamental and applied studies. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Automated magnification calibration in transmission electron microscopy using Fourier analysis of replica images.

PubMed

van der Laak, Jeroen A W M; Dijkman, Henry B P M; Pahlplatz, Martin M M

2006-03-01

The magnification factor in transmission electron microscopy is not very precise, hampering for instance quantitative analysis of specimens. Calibration of the magnification is usually performed interactively using replica specimens, containing line or grating patterns with known spacing. In the present study, a procedure is described for automated magnification calibration using digital images of a line replica. This procedure is based on analysis of the power spectrum of Fourier transformed replica images, and is compared to interactive measurement in the same images. Images were used with magnification ranging from 1,000 x to 200,000 x. The automated procedure deviated on average 0.10% from interactive measurements. Especially for catalase replicas, the coefficient of variation of automated measurement was considerably smaller (average 0.28%) compared to that of interactive measurement (average 3.5%). In conclusion, calibration of the magnification in digital images from transmission electron microscopy may be performed automatically, using the procedure presented here, with high precision and accuracy.

Methods of Hematoxylin and Erosin Image Information Acquisition and Optimization in Confocal Microscopy

PubMed Central

Yoon, Woong Bae; Kim, Hyunjin; Kim, Kwang Gi; Choi, Yongdoo; Chang, Hee Jin

2016-01-01

Objectives We produced hematoxylin and eosin (H&E) staining-like color images by using confocal laser scanning microscopy (CLSM), which can obtain the same or more information in comparison to conventional tissue staining. Methods We improved images by using several image converting techniques, including morphological methods, color space conversion methods, and segmentation methods. Results An image obtained after image processing showed coloring very similar to that in images produced by H&E staining, and it is advantageous to conduct analysis through fluorescent dye imaging and microscopy rather than analysis based on single microscopic imaging. Conclusions The colors used in CLSM are different from those seen in H&E staining, which is the method most widely used for pathologic diagnosis and is familiar to pathologists. Computer technology can facilitate the conversion of images by CLSM to be very similar to H&E staining images. We believe that the technique used in this study has great potential for application in clinical tissue analysis. PMID:27525165

Robust analysis method for acoustic properties of biological specimens measured by acoustic microscopy

NASA Astrophysics Data System (ADS)

Arakawa, Mototaka; Mori, Shohei; Kanai, Hiroshi; Nagaoka, Ryo; Horie, Miki; Kobayashi, Kazuto; Saijo, Yoshifumi

2018-07-01

We proposed a robust analysis method for the acoustic properties of biological specimens measured by acoustic microscopy. Reflected pulse signals from the substrate and specimen were converted into frequency domains to obtain sound speed and thickness. To obtain the average acoustic properties of the specimen, parabolic approximation was performed to determine the frequency at which the amplitude of the normalized spectrum became maximum or minimum, considering the sound speed and thickness of the specimens and the operating frequency of the ultrasonic device used. The proposed method was demonstrated for a specimen of malignant melanoma of the skin by using acoustic microscopy attaching a concave transducer with a center frequency of 80 MHz. The variations in sound speed and thickness analyzed by the proposed method were markedly smaller than those analyzed by the method based on an autoregressive model. The proposed method is useful for the analysis of the acoustic properties of bilogical tissues or cells.

A coarse-grained model for DNA origami.

PubMed

Reshetnikov, Roman V; Stolyarova, Anastasia V; Zalevsky, Arthur O; Panteleev, Dmitry Y; Pavlova, Galina V; Klinov, Dmitry V; Golovin, Andrey V; Protopopova, Anna D

2018-02-16

Modeling tools provide a valuable support for DNA origami design. However, current solutions have limited application for conformational analysis of the designs. In this work we present a tool for a thorough study of DNA origami structure and dynamics. The tool is based on a novel coarse-grained model dedicated to geometry optimization and conformational analysis of DNA origami. We explored the ability of the model to predict dynamic behavior, global shapes, and fine details of two single-layer systems designed in hexagonal and square lattices using atomic force microscopy, Förster resonance energy transfer spectroscopy, and all-atom molecular dynamic simulations for validation of the results. We also examined the performance of the model for multilayer systems by simulation of DNA origami with published cryo-electron microscopy and atomic force microscopy structures. A good agreement between the simulated and experimental data makes the model suitable for conformational analysis of DNA origami objects. The tool is available at http://vsb.fbb.msu.ru/cosm as a web-service and as a standalone version.

Characterization of Deposits on Glass Substrate as a Tool in Failure Analysis: The Orbiter Vehicle Columbia Case Study

NASA Technical Reports Server (NTRS)

Olivas, J. D.; Melroy, P.; McDanels, S.; Wallace, T.; Zapata, M. C.

2006-01-01

In connection with the accident investigation of the space shuttle Columbia, an analysis methodology utilizing well established microscopic and spectroscopic techniques was implemented for evaluating the environment to which the exterior fused silica glass was exposed. Through the implementation of optical microscopy, scanning electron microscopy, energy dispersive spectroscopy, transmission electron microscopy, and electron diffraction, details emerged regarding the manner in which a charred metallic deposited layer formed on top of the exposed glass. Due to nature of the substrate and the materials deposited, the methodology proved to allow for a more detailed analysis of the vehicle breakup. By contrast, similar analytical methodologies on metallic substrates have proven to be challenging due to strong potential for error resulting from substrate contamination. This information proved to be valuable to not only those involved in investigating the break up of Columbia, but also provides a potential guide for investigating future high altitude and high energy accidents.

Towards a nondestructive chemical characterization of biofilm matrix by Raman microscopy.

PubMed

Ivleva, Natalia P; Wagner, Michael; Horn, Harald; Niessner, Reinhard; Haisch, Christoph

2009-01-01

In this study, the applicability of Raman microscopy (RM) for nondestructive chemical analysis of biofilm matrix, including microbial constituents and extracellular polymeric substances (EPS), has been assessed. The examination of a wide range of reference samples such as biofilm-specific polysaccharides, proteins, microorganisms, and encapsulated bacteria revealed characteristic frequency regions and specific marker bands for different biofilm constituents. Based on received data, the assignment of Raman bands in spectra of multispecies biofilms was performed. The study of different multispecies biofilms showed that RM can correlate various structural appearances within the biofilm to variations in their chemical composition and provide chemical information about a complex biofilm matrix. The results of RM analysis of biofilms are in good agreement with data obtained by confocal laser scanning microscopy (CLSM). Thus, RM is a promising tool for a label-free chemical characterization of different biofilm constituents. Moreover, the combination of RM with CLSM analysis for the study of biofilms grown under different environmental conditions can provide new insights into the complex structure/function correlations in biofilms.

Methods of Hematoxylin and Erosin Image Information Acquisition and Optimization in Confocal Microscopy.

PubMed

Yoon, Woong Bae; Kim, Hyunjin; Kim, Kwang Gi; Choi, Yongdoo; Chang, Hee Jin; Sohn, Dae Kyung

2016-07-01

We produced hematoxylin and eosin (H&E) staining-like color images by using confocal laser scanning microscopy (CLSM), which can obtain the same or more information in comparison to conventional tissue staining. We improved images by using several image converting techniques, including morphological methods, color space conversion methods, and segmentation methods. An image obtained after image processing showed coloring very similar to that in images produced by H&E staining, and it is advantageous to conduct analysis through fluorescent dye imaging and microscopy rather than analysis based on single microscopic imaging. The colors used in CLSM are different from those seen in H&E staining, which is the method most widely used for pathologic diagnosis and is familiar to pathologists. Computer technology can facilitate the conversion of images by CLSM to be very similar to H&E staining images. We believe that the technique used in this study has great potential for application in clinical tissue analysis.

A coarse-grained model for DNA origami

PubMed Central

Stolyarova, Anastasia V; Zalevsky, Arthur O; Panteleev, Dmitry Y; Pavlova, Galina V; Klinov, Dmitry V; Golovin, Andrey V; Protopopova, Anna D

2018-01-01

Abstract Modeling tools provide a valuable support for DNA origami design. However, current solutions have limited application for conformational analysis of the designs. In this work we present a tool for a thorough study of DNA origami structure and dynamics. The tool is based on a novel coarse-grained model dedicated to geometry optimization and conformational analysis of DNA origami. We explored the ability of the model to predict dynamic behavior, global shapes, and fine details of two single-layer systems designed in hexagonal and square lattices using atomic force microscopy, Förster resonance energy transfer spectroscopy, and all-atom molecular dynamic simulations for validation of the results. We also examined the performance of the model for multilayer systems by simulation of DNA origami with published cryo-electron microscopy and atomic force microscopy structures. A good agreement between the simulated and experimental data makes the model suitable for conformational analysis of DNA origami objects. The tool is available at http://vsb.fbb.msu.ru/cosm as a web-service and as a standalone version. PMID:29267876

A simple approach to characterizing block copolymer assemblies: graphene oxide supports for high contrast multi-technique imaging†

PubMed Central

Patterson, Joseph P.; Sanchez, Ana M.; Petzetakis, Nikos; Smart, Thomas P.; Epps, Thomas H.; Portman, Ian

2013-01-01

Block copolymers are well-known to self-assemble into a range of 3-dimensional morphologies. However, due to their nanoscale dimensions, resolving their exact structure can be a challenge. Transmission electron microscopy (TEM) is a powerful technique for achieving this, but for polymeric assemblies chemical fixing/staining techniques are usually required to increase image contrast and protect specimens from electron beam damage. Graphene oxide (GO) is a robust, water-dispersable, and nearly electron transparent membrane: an ideal support for TEM. We show that when using GO supports no stains are required to acquire high contrast TEM images and that the specimens remain stable under the electron beam for long periods, allowing sample analysis by a range of electron microscopy techniques. GO supports are also used for further characterization of assemblies by atomic force microscopy. The simplicity of sample preparation and analysis, as well as the potential for significantly increased contrast background, make GO supports an attractive alternative for the analysis of block copolymer assemblies. PMID:24049544

Accounting for Limited Detection Efficiency and Localization Precision in Cluster Analysis in Single Molecule Localization Microscopy

PubMed Central

Shivanandan, Arun; Unnikrishnan, Jayakrishnan; Radenovic, Aleksandra

2015-01-01

Single Molecule Localization Microscopy techniques like PhotoActivated Localization Microscopy, with their sub-diffraction limit spatial resolution, have been popularly used to characterize the spatial organization of membrane proteins, by means of quantitative cluster analysis. However, such quantitative studies remain challenged by the techniques’ inherent sources of errors such as a limited detection efficiency of less than 60%, due to incomplete photo-conversion, and a limited localization precision in the range of 10 – 30nm, varying across the detected molecules, mainly depending on the number of photons collected from each. We provide analytical methods to estimate the effect of these errors in cluster analysis and to correct for them. These methods, based on the Ripley’s L(r) – r or Pair Correlation Function popularly used by the community, can facilitate potentially breakthrough results in quantitative biology by providing a more accurate and precise quantification of protein spatial organization. PMID:25794150

Standardization for Ki-67 Assessment in Moderately Differentiated Breast Cancer. A Retrospective Analysis of the SAKK 28/12 Study

PubMed Central

Varga, Zsuzsanna; Cassoly, Estelle; Li, Qiyu; Oehlschlegel, Christian; Tapia, Coya; Lehr, Hans Anton; Klingbiel, Dirk; Thürlimann, Beat; Ruhstaller, Thomas

2015-01-01

Background Proliferative activity (Ki-67 Labelling Index) in breast cancer increasingly serves as an additional tool in the decision for or against adjuvant chemotherapy in midrange hormone receptor positive breast cancer. Ki-67 Index has been previously shown to suffer from high inter-observer variability especially in midrange (G2) breast carcinomas. In this study we conducted a systematic approach using different Ki-67 assessments on large tissue sections in order to identify the method with the highest reliability and the lowest variability. Materials and Methods Five breast pathologists retrospectively analyzed proliferative activity of 50 G2 invasive breast carcinomas using large tissue sections by assessing Ki-67 immunohistochemistry. Ki-67-assessments were done on light microscopy and on digital images following these methods: 1) assessing five regions, 2) assessing only darkly stained nuclei and 3) considering only condensed proliferative areas (‘hotspots’). An individual review (the first described assessment from 2008) was also performed. The assessments on light microscopy were done by estimating. All measurements were performed three times. Inter-observer and intra-observer reliabilities were calculated using the approach proposed by Eliasziw et al. Clinical cutoffs (14% and 20%) were tested using Fleiss’ Kappa. Results There was a good intra-observer reliability in 5 of 7 methods (ICC: 0.76–0.89). The two highest inter-observer reliability was fair to moderate (ICC: 0.71 and 0.74) in 2 methods (region-analysis and individual-review) on light microscopy. Fleiss’-kappa-values (14% cut-off) were the highest (moderate) using the original recommendation on light-microscope (Kappa 0.58). Fleiss’ kappa values (20% cut-off) were the highest (Kappa 0.48 each) in analyzing hotspots on light-microscopy and digital-analysis. No methodologies using digital-analysis were superior to the methods on light microscope. Conclusion Our results show that all methods on light-microscopy for Ki-67 assessment in large tissue sections resulted in a good intra-observer reliability. Region analysis and individual review (the original recommendation) on light-microscopy yielded the highest inter-observer reliability. These results show slight improvement to previously published data on poor-reproducibility and thus might be a practical-pragmatic way for routine assessment of Ki-67 Index in G2 breast carcinomas. PMID:25885288

A Bayesian cluster analysis method for single-molecule localization microscopy data.

PubMed

Griffié, Juliette; Shannon, Michael; Bromley, Claire L; Boelen, Lies; Burn, Garth L; Williamson, David J; Heard, Nicholas A; Cope, Andrew P; Owen, Dylan M; Rubin-Delanchy, Patrick

2016-12-01

Cell function is regulated by the spatiotemporal organization of the signaling machinery, and a key facet of this is molecular clustering. Here, we present a protocol for the analysis of clustering in data generated by 2D single-molecule localization microscopy (SMLM)-for example, photoactivated localization microscopy (PALM) or stochastic optical reconstruction microscopy (STORM). Three features of such data can cause standard cluster analysis approaches to be ineffective: (i) the data take the form of a list of points rather than a pixel array; (ii) there is a non-negligible unclustered background density of points that must be accounted for; and (iii) each localization has an associated uncertainty in regard to its position. These issues are overcome using a Bayesian, model-based approach. Many possible cluster configurations are proposed and scored against a generative model, which assumes Gaussian clusters overlaid on a completely spatially random (CSR) background, before every point is scrambled by its localization precision. We present the process of generating simulated and experimental data that are suitable to our algorithm, the analysis itself, and the extraction and interpretation of key cluster descriptors such as the number of clusters, cluster radii and the number of localizations per cluster. Variations in these descriptors can be interpreted as arising from changes in the organization of the cellular nanoarchitecture. The protocol requires no specific programming ability, and the processing time for one data set, typically containing 30 regions of interest, is ∼18 h; user input takes ∼1 h.

Tracking molecular dynamics without tracking: image correlation of photo-activation microscopy

NASA Astrophysics Data System (ADS)

Pandžić, Elvis; Rossy, Jérémie; Gaus, Katharina

2015-03-01

Measuring protein dynamics in the plasma membrane can provide insights into the mechanisms of receptor signaling and other cellular functions. To quantify protein dynamics on the single molecule level over the entire cell surface, sophisticated approaches such as single particle tracking (SPT), photo-activation localization microscopy (PALM) and fluctuation-based analysis have been developed. However, analyzing molecular dynamics of fluorescent particles with intermittent excitation and low signal-to-noise ratio present at high densities has remained a challenge. We overcame this problem by applying spatio-temporal image correlation spectroscopy (STICS) analysis to photo-activated (PA) microscopy time series. In order to determine under which imaging conditions this approach is valid, we simulated PA images of diffusing particles in a homogeneous environment and varied photo-activation, reversible blinking and irreversible photo-bleaching rates. Further, we simulated data with high particle densities that populated mobile objects (such as adhesions and vesicles) that often interfere with STICS and fluctuation-based analysis. We demonstrated in experimental measurements that the diffusion coefficient of the epidermal growth factor receptor (EGFR) fused to PAGFP in live COS-7 cells can be determined in the plasma membrane and revealed differences in the time-dependent diffusion maps between wild-type and mutant Lck in activated T cells. In summary, we have developed a new analysis approach for live cell photo-activation microscopy data based on image correlation spectroscopy to quantify the spatio-temporal dynamics of single proteins.

Simulation of bright-field microscopy images depicting pap-smear specimen

PubMed Central

Malm, Patrik; Brun, Anders; Bengtsson, Ewert

2015-01-01

As digital imaging is becoming a fundamental part of medical and biomedical research, the demand for computer-based evaluation using advanced image analysis is becoming an integral part of many research projects. A common problem when developing new image analysis algorithms is the need of large datasets with ground truth on which the algorithms can be tested and optimized. Generating such datasets is often tedious and introduces subjectivity and interindividual and intraindividual variations. An alternative to manually created ground-truth data is to generate synthetic images where the ground truth is known. The challenge then is to make the images sufficiently similar to the real ones to be useful in algorithm development. One of the first and most widely studied medical image analysis tasks is to automate screening for cervical cancer through Pap-smear analysis. As part of an effort to develop a new generation cervical cancer screening system, we have developed a framework for the creation of realistic synthetic bright-field microscopy images that can be used for algorithm development and benchmarking. The resulting framework has been assessed through a visual evaluation by experts with extensive experience of Pap-smear images. The results show that images produced using our described methods are realistic enough to be mistaken for real microscopy images. The developed simulation framework is very flexible and can be modified to mimic many other types of bright-field microscopy images. © 2015 The Authors. Published by Wiley Periodicals, Inc. on behalf of ISAC PMID:25573002

Discovery: Under the Microscope at Kennedy Space Center

NASA Technical Reports Server (NTRS)

Howard, Philip M.

2013-01-01

The National Aeronautics & Space Administration (NASA) is known for discovery, exploration, and advancement of knowledge. Since the days of Leeuwenhoek, microscopy has been at the forefront of discovery and knowledge. No truer is that statement than today at Kennedy Space Center (KSC), where microscopy plays a major role in contamination identification and is an integral part of failure analysis. Space exploration involves flight hardware undergoing rigorous "visually clean" inspections at every step of processing. The unknown contaminants that are discovered on these inspections can directly impact the mission by decreasing performance of sensors and scientific detectors on spacecraft and satellites, acting as micrometeorites, damaging critical sealing surfaces, and causing hazards to the crew of manned missions. This talk will discuss how microscopy has played a major role in all aspects of space port operations at KSC. Case studies will highlight years of analysis at the Materials Science Division including facility and payload contamination for the Navigation Signal Timing and Ranging Global Positioning Satellites (NA VST AR GPS) missions, quality control monitoring of monomethyl hydrazine fuel procurement for launch vehicle operations, Shuttle Solids Rocket Booster (SRB) foam processing failure analysis, and Space Shuttle Main Engine Cut-off (ECO) flight sensor anomaly analysis. What I hope to share with my fellow microscopists is some of the excitement of microscopy and how its discoveries has led to hardware processing, that has helped enable the successful launch of vehicles and space flight missions here at Kennedy Space Center.

Tracking molecular dynamics without tracking: image correlation of photo-activation microscopy.

PubMed

Pandžić, Elvis; Rossy, Jérémie; Gaus, Katharina

2015-03-09

Measuring protein dynamics in the plasma membrane can provide insights into the mechanisms of receptor signaling and other cellular functions. To quantify protein dynamics on the single molecule level over the entire cell surface, sophisticated approaches such as single particle tracking (SPT), photo-activation localization microscopy (PALM) and fluctuation-based analysis have been developed. However, analyzing molecular dynamics of fluorescent particles with intermittent excitation and low signal-to-noise ratio present at high densities has remained a challenge. We overcame this problem by applying spatio-temporal image correlation spectroscopy (STICS) analysis to photo-activated (PA) microscopy time series. In order to determine under which imaging conditions this approach is valid, we simulated PA images of diffusing particles in a homogeneous environment and varied photo-activation, reversible blinking and irreversible photo-bleaching rates. Further, we simulated data with high particle densities that populated mobile objects (such as adhesions and vesicles) that often interfere with STICS and fluctuation-based analysis. We demonstrated in experimental measurements that the diffusion coefficient of the epidermal growth factor receptor (EGFR) fused to PAGFP in live COS-7 cells can be determined in the plasma membrane and revealed differences in the time-dependent diffusion maps between wild-type and mutant Lck in activated T cells. In summary, we have developed a new analysis approach for live cell photo-activation microscopy data based on image correlation spectroscopy to quantify the spatio-temporal dynamics of single proteins.

Image improvement and three-dimensional reconstruction using holographic image processing

NASA Technical Reports Server (NTRS)

Stroke, G. W.; Halioua, M.; Thon, F.; Willasch, D. H.

1977-01-01

Holographic computing principles make possible image improvement and synthesis in many cases of current scientific and engineering interest. Examples are given for the improvement of resolution in electron microscopy and 3-D reconstruction in electron microscopy and X-ray crystallography, following an analysis of optical versus digital computing in such applications.

CHEMICAL MAPPING OF ELEMENTAL SULFUR ON PYRITE AND ARSENOPYRITE SURFACES USING NEAR-INFRARED RAMAN IMAGING MICROSCOPY. (R826189)

EPA Science Inventory

Abstract

Near-infrared Raman imaging microscopy (NIRIM) was used to produce chemical images of the distribution of elemental sulfur on oxidized pyrite and arsenopyrite surfaces. Analysis using Savitsky_¯Golay filtering permits an unambiguous identificati...

Analytical Microscopy and Imaging Science | Materials Science | NREL

Science.gov Websites

Microanalysis (EPMA) for quantitative compositional analysis. It relies on wavelength-dispersive spectroscopy to Science group in NREL's Materials Science Center. Mowafak Al-Jassim Group Manager Dr. Al-Jassim manages the Analytical Microscopy and Imaging Science group with the Materials Science Center. Email | 303-384

Comparative study of three Plumbago L. species (Plumbaginaceae) by microscopy, UPLC–UV and HPTLC analyses

USDA-ARS?s Scientific Manuscript database

This paper presents a comparative study of anatomy of leaves, stems and roots of three species of Plumbago, namely P. auriculata Lam., P. indica L. and P. zeylanica L. by light microscopy. The paper also provides qualitative and quantitative analysis of the naphthoquinone, plumbagin, a major constit...

Correlative Microscopy of Vitreous Sections Provides Insights into BAR-Domain Organization In Situ.

PubMed

Bharat, Tanmay A M; Hoffmann, Patrick C; Kukulski, Wanda

2018-04-10

Electron microscopy imaging of macromolecular complexes in their native cellular context is limited by the inherent difficulty to acquire high-resolution tomographic data from thick cells and to specifically identify elusive structures within crowded cellular environments. Here, we combined cryo-fluorescence microscopy with electron cryo-tomography of vitreous sections into a coherent correlative microscopy workflow, ideal for detection and structural analysis of elusive protein assemblies in situ. We used this workflow to address an open question on BAR-domain coating of yeast plasma membrane compartments known as eisosomes. BAR domains can sense or induce membrane curvature, and form scaffold-like membrane coats in vitro. Our results demonstrate that in cells, the BAR protein Pil1 localizes to eisosomes of varying membrane curvature. Sub-tomogram analysis revealed a dense protein coat on curved eisosomes, which was not present on shallow eisosomes, indicating that while BAR domains can assemble at shallow membranes in vivo, scaffold formation is tightly coupled to curvature generation. Copyright © 2018 MRC Laboratory of Molecular Biology. Published by Elsevier Ltd.. All rights reserved.

Scanning fluorescent microscopy is an alternative for quantitative fluorescent cell analysis.

PubMed

Varga, Viktor Sebestyén; Bocsi, József; Sipos, Ferenc; Csendes, Gábor; Tulassay, Zsolt; Molnár, Béla

2004-07-01

Fluorescent measurements on cells are performed today with FCM and laser scanning cytometry. The scientific community dealing with quantitative cell analysis would benefit from the development of a new digital multichannel and virtual microscopy based scanning fluorescent microscopy technology and from its evaluation on routine standardized fluorescent beads and clinical specimens. We applied a commercial motorized fluorescent microscope system. The scanning was done at 20 x (0.5 NA) magnification, on three channels (Rhodamine, FITC, Hoechst). The SFM (scanning fluorescent microscopy) software included the following features: scanning area, exposure time, and channel definition, autofocused scanning, densitometric and morphometric cellular feature determination, gating on scatterplots and frequency histograms, and preparation of galleries of the gated cells. For the calibration and standardization Immuno-Brite beads were used. With application of shading compensation, the CV of fluorescence of the beads decreased from 24.3% to 3.9%. Standard JPEG image compression until 1:150 resulted in no significant change. The change of focus influenced the CV significantly only after +/-5 microm error. SFM is a valuable method for the evaluation of fluorescently labeled cells. Copyright 2004 Wiley-Liss, Inc.

Experiments in electron microscopy: from metals to nerves

NASA Astrophysics Data System (ADS)

Unwin, Nigel

2015-04-01

Electron microscopy has advanced remarkably as a tool for biological structure research since the development of methods to examine radiation-sensitive unstained specimens and the introduction of cryo-techniques. Structures of biological molecules at near-atomic resolution can now be obtained from images of single particles as well as crystalline arrays. It has also become possible to analyze structures of molecules in their functional context, i.e. in their natural membrane or cellular setting, and in an ionic environment like that in living tissue. Electron microscopy is thus opening ways to answer definitively questions about physiological mechanisms. Here I recall a number of experiments contributing to, and benefiting from the technical advances that have taken place. I begin—in the spirit of this crystallography series—with some biographical background, and then sketch the path to an analysis by time-resolved microscopy of the opening mechanism of an ion channel (nicotinic acetylcholine receptor). This analysis illustrates how electron imaging can be combined with freeze-trapping to illuminate a transient biological event: in our case, chemical-to-electrical transduction at the nerve-muscle synapse.

Technician Consistency in Specular Microscopy Measurements: A "Real-World" Retrospective Analysis of a United States Eye Bank.

PubMed

Rand, Gabriel M; Kwon, Ji Won; Gore, Patrick K; McCartney, Mitchell D; Chuck, Roy S

2017-10-01

To quantify consistency of endothelial cell density (ECD) measurements among technicians in a single US eye bank operating under typical operating conditions. In this retrospective analysis of 51 microscopy technicians using a semiautomated counting method on 35,067 eyes from July 2007 to May 2015, technician- and date-related marginal ECD effects were calculated using linear regression models. ECD variance was correlated with the number of specular microscopy technicians. Technician mean ECDs ranged from 2386 ± 431 to 3005 ± 560 cells/mm. Nine technicians had statistically and clinically significant marginal effects. Annual mean ECDs adjusted for changes in technicians ranged from 2422 ± 433 to 2644 ± 430 cells/mm. The period of 2007 to 2009 had statistically and clinically significant marginal effects. There was a nonstatistically significant association between the number of technicians and ECD standard deviation. There was significant ECD variability associated with specular microscopy technicians and with the date of measurement. We recommend that eye banks collect data related to laboratory factors that have been shown to influence ECD variability.

Scanning Tunneling Microscopy analysis of space-exposed polymer films

NASA Technical Reports Server (NTRS)

Kalil, Carol R.; Young, Philip R.

1993-01-01

The characterization of the surface of selected space-exposed polymer films by Scanning Tunneling Microscopy (STM) is reported. Principles of STM, an emerging new technique for materials analysis, are reviewed. The analysis of several films which received up to 5.8 years of low Earth orbital (LEO) exposure onboard the NASA Long Duration Exposure Facility (LDEF) is discussed. Specimens included FEP Teflon thermal blanket material, Kapton film, and several experimental polymer films. Ultraviolet and atomic oxygen-induced crazing and erosion are described. The intent of this paper is to demonstrate how STM is enhancing the understanding of LEO space environmental effects on polymer films.

Spectrometric analysis and scanning electronic microscopy of two pleural plaques from mediaeval Portuguese period.

PubMed

Fernandes, T; Granja, R; Thillaud, P L

2014-01-01

During an archaeological excavation at a mediaeval monastery (Flor da Rosa, Crato, Portugal), a skeleton of a adult woman was found with two calcifications in the thoracic cage. The location and the macroscopic analysis of the calcifications allowed them to be assigned as pleural plaques. Spectrometric analysis and scanning electronic microscopy enabled to establish that it originated with an infectious process. These results associated with the lesions found in the ribs and vertebrae strongly suggest tuberculosis as the cause of these pleural plaques. Copyright © 2013 Sociedade Portuguesa de Pneumologia. Published by Elsevier España. All rights reserved.

X-ray microscopy of live biological micro-organisms

NASA Astrophysics Data System (ADS)

Raja Al-Ani, Ma'an Nassar

Real-time, compact x-ray microscopy has the potential to benefit many scientific fields, including microbiology, pharmacology, organic chemistry, and physics. Single frame x-ray micro-radiography, produced by a compact, solid-state laser plasma source, allows scientists to use x-ray emission for elemental analysis, and to observe biological specimens in their natural state. In this study, x-ray images of mouse kidney tissue, live bacteria, Pseudomonas aeruginosa and Burkholderia cepacia, and the bacteria's interaction with the antibiotic gentamicin, are examined using x-ray microscopy. For the purposes of comparing between confocal microscopy and x-ray microscopy, we introduced to our work the technique of gold labeling. Indirect immunofluorescence staining and immuno-gold labeling were applied on human lymphocytes and human tumor cells. Differential interference contrast microscopy (DIC) showed the lymphocyte body and nucleus, as did x-ray microscopy. However, the high resolution of x-ray microscopy allows us to differentiate between the gold particles bound to the antibodies and the free gold. A compact, tabletop Nd: glass laser is used in this study to produce x-rays from an Yttrium target. An atomic force microscope is used to scan the x-ray images from the developed photo-resist. The use of compact, tabletop laser plasma sources, in conjunction with x-ray microscopy, is a new technique that has great potential as a flexible, user-friendly scientific research tool.

Simulated single molecule microscopy with SMeagol.

PubMed

Lindén, Martin; Ćurić, Vladimir; Boucharin, Alexis; Fange, David; Elf, Johan

2016-08-01

SMeagol is a software tool to simulate highly realistic microscopy data based on spatial systems biology models, in order to facilitate development, validation and optimization of advanced analysis methods for live cell single molecule microscopy data. SMeagol runs on Matlab R2014 and later, and uses compiled binaries in C for reaction-diffusion simulations. Documentation, source code and binaries for Mac OS, Windows and Ubuntu Linux can be downloaded from http://smeagol.sourceforge.net johan.elf@icm.uu.se Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press.

Correlative light-electron fractography for fatigue striations characterization in metallic alloys.

PubMed

Hein, Luis Rogerio de Oliveira; de Oliveira, José Alberto; de Campos, Kamila Amato

2013-09-01

The correlative light-electron fractography technique combines correlative microscopy concepts to the extended depth-from-focus reconstruction method, associating the reliable topographic information of 3-D maps from light microscopy ordered Z-stacks to the finest lateral resolution and large focus depth from scanning electron microscopy. Fatigue striations spacing analysis can be precisely measured, by correcting the mean surface tilting with the knowledge of local elevation data from elevation maps. This new technique aims to improve the accuracy of quantitative fractography in fatigue fracture investigations. Copyright © 2013 Wiley Periodicals, Inc.

Transmission Electron Microscopy Analysis of Skin Lesions from Sporotrichosis Epidemic in Rio de Janeiro, Brazil

PubMed Central

Porto Ferreira, Cassio; Oliveira de Almeida, Ana Cristina; Corte-Real, Suzana

2015-01-01

Transmission electron microscopy can yield useful information in a range of scientific fields; it is capable of imaging at a significantly higher resolution than light microscopes and has been a very useful tool in the identification of morphological changes of the dermis as well as assessment of changes in the extracellular matrix. Our aim is to characterize by electron microscopy the cellular profile of lesions caused by Sporothrix schenckii from the sporotrichosis epidemic in its zoonotic form that occurs in Rio de Janeiro, Brazil. PMID:25653392

Nanoscale quantification of intracellular element concentration by X-ray fluorescence microscopy combined with X-ray phase contrast nanotomography

NASA Astrophysics Data System (ADS)

Gramaccioni, Chiara; Yang, Yang; Procopio, Alessandra; Pacureanu, Alexandra; Bohic, Sylvain; Malucelli, Emil; Iotti, Stefano; Farruggia, Giovanna; Bukreeva, Inna; Notargiacomo, Andrea; Fratini, Michela; Valenti, Piera; Rosa, Luigi; Berlutti, Francesca; Cloetens, Peter; Lagomarsino, Stefano

2018-01-01

We present here a correlative X-ray microscopy approach for quantitative single cell imaging of molar concentrations. By combining the elemental content provided by X-ray fluorescence microscopy and the morphology information extracted from X-ray phase nanotomography, we determine the intracellular molarity distributions. This correlative method was demonstrated on a freeze-dried human phagocytic cell to obtain the absolute elemental concentration maps of K, P, and Fe. The cell morphology results showed a very good agreement with atomic-force microscopy measurements. This work opens the way for non-destructive single cell chemical analysis down to the sub-cellular level using exclusively synchrotron radiation techniques. It will be of high interest in the case where it is difficult to access the morphology using atomic-force microscopy, for example, on frozen-hydrated cells or tissues.

Correlative microscopy of detergent granules.

PubMed

van Dalen, G; Nootenboom, P; Heussen, P C M

2011-03-01

The microstructure of detergent products for textile cleaning determines to a large extent the physical properties of these products. Correlative microscopy was used to reveal the microstructure by reconciling images obtained by scanning electron microscopy with energy dispersive X-ray analysis, X-ray microtomography and Fourier transform infrared microscopy. These techniques were applied on the same location of a subsample of a spray-dried detergent base powder embedded in polyacrylate. In this way, the three-dimensional internal and external structure of detergent granules could be investigated from milli to nano scale with detailed spatial information about the components present. This will generate knowledge how to design optimal microstructures for laundry products to obtain product properties demanded by the market. This method is also very useful for other powder systems used in a large variety of industries (e.g. for pharmaceutical, food, ceramic and metal industries). © 2010 The Authors Journal of Microscopy © 2010 The Royal Microscopical Society.

Hybrid microscopy of human carotid atheroma by means of optical-resolution optoacoustic and non-linear optical microscopy

NASA Astrophysics Data System (ADS)

Seeger, Markus; Karlas, Angelos; Soliman, Dominik; Pelisek, Jaroslav; Ntziachristos, Vasilis

2017-03-01

Carotid atheromatosis is causally related to stroke, a leading cause of disability and death. We present the analysis of a human carotid atheroma using a novel hybrid microscopy system that combines optical-resolution optoacoustic (photoacoustic) microscopy and several non-linear optical microscopy modalities (second and third harmonic generation, as well as, two-photon excitation fluorescence) to achieve a multimodal examination of the extracted tissue within the same imaging framework. Our system enables the label-free investigation of atheromatous human carotid tissue with a resolution of about 1 μm and allows for the congruent interrogation of plaque morphology and clinically relevant constituents such as red blood cells, collagen, and elastin. Our data reveal mutual interactions between blood embeddings and connective tissue within the atheroma, offering comprehensive insights into its stage of evolution and severity, and potentially facilitating the further development of diagnostic tools, as well as treatment strategies.

Nanoscale surface characterization using laser interference microscopy

NASA Astrophysics Data System (ADS)

Ignatyev, Pavel S.; Skrynnik, Andrey A.; Melnik, Yury A.

2018-03-01

Nanoscale surface characterization is one of the most significant parts of modern materials development and application. The modern microscopes are expensive and complicated tools, and its use for industrial tasks is limited due to laborious sample preparation, measurement procedures, and low operation speed. The laser modulation interference microscopy method (MIM) for real-time quantitative and qualitative analysis of glass, metals, ceramics, and various coatings has a spatial resolution of 0.1 nm for vertical and up to 100 nm for lateral. It is proposed as an alternative to traditional scanning electron microscopy (SEM) and atomic force microscopy (AFM) methods. It is demonstrated that in the cases of roughness metrology for super smooth (Ra >1 nm) surfaces the application of a laser interference microscopy techniques is more optimal than conventional SEM and AFM. The comparison of semiconductor test structure for lateral dimensions measurements obtained with SEM and AFM and white light interferometer also demonstrates the advantages of MIM technique.

SpectraFox: A free open-source data management and analysis tool for scanning probe microscopy and spectroscopy

NASA Astrophysics Data System (ADS)

Ruby, Michael

In the last decades scanning probe microscopy and spectroscopy have become well-established tools in nanotechnology and surface science. This opened the market for many commercial manufacturers, each with different hardware and software standards. Besides the advantage of a wide variety of available hardware, the diversity may software-wise complicate the data exchange between scientists, and the data analysis for groups working with hardware developed by different manufacturers. Not only the file format differs between manufacturers, but also the data often requires further numerical treatment before publication. SpectraFox is an open-source and independent tool which manages, processes, and evaluates scanning probe spectroscopy and microscopy data. It aims at simplifying the documentation in parallel to measurement, and it provides solid evaluation tools for a large number of data.

Automated quantitative cytological analysis using portable microfluidic microscopy.

PubMed

Jagannadh, Veerendra Kalyan; Murthy, Rashmi Sreeramachandra; Srinivasan, Rajesh; Gorthi, Sai Siva

2016-06-01

In this article, a portable microfluidic microscopy based approach for automated cytological investigations is presented. Inexpensive optical and electronic components have been used to construct a simple microfluidic microscopy system. In contrast to the conventional slide-based methods, the presented method employs microfluidics to enable automated sample handling and image acquisition. The approach involves the use of simple in-suspension staining and automated image acquisition to enable quantitative cytological analysis of samples. The applicability of the presented approach to research in cellular biology is shown by performing an automated cell viability assessment on a given population of yeast cells. Further, the relevance of the presented approach to clinical diagnosis and prognosis has been demonstrated by performing detection and differential assessment of malaria infection in a given sample. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Live cell refractometry using Hilbert phase microscopy and confocal reflectance microscopy.

PubMed

Lue, Niyom; Choi, Wonshik; Popescu, Gabriel; Yaqoob, Zahid; Badizadegan, Kamran; Dasari, Ramachandra R; Feld, Michael S

2009-11-26

Quantitative chemical analysis has served as a useful tool for understanding cellular metabolisms in biology. Among many physical properties used in chemical analysis, refractive index in particular has provided molecular concentration that is an important indicator for biological activities. In this report, we present a method of extracting full-field refractive index maps of live cells in their native states. We first record full-field optical thickness maps of living cells by Hilbert phase microscopy and then acquire physical thickness maps of the same cells using a custom-built confocal reflectance microscope. Full-field and axially averaged refractive index maps are acquired from the ratio of optical thickness to physical thickness. The accuracy of the axially averaged index measurement is 0.002. This approach can provide novel biological assays of label-free living cells in situ.

Correlative fractography: combining scanning electron microscopy and light microscopes for qualitative and quantitative analysis of fracture surfaces.

PubMed

Hein, Luis Rogerio de Oliveira; de Oliveira, José Alberto; de Campos, Kamila Amato

2013-04-01

Correlative fractography is a new expression proposed here to describe a new method for the association between scanning electron microscopy (SEM) and light microscopy (LM) for the qualitative and quantitative analysis of fracture surfaces. This article presents a new method involving the fusion of one elevation map obtained by extended depth from focus reconstruction from LM with exactly the same area by SEM and associated techniques, as X-ray mapping. The true topographic information is perfectly associated to local fracture mechanisms with this new technique, presented here as an alternative to stereo-pair reconstruction for the investigation of fractured components. The great advantage of this technique resides in the possibility of combining any imaging methods associated with LM and SEM for the same observed field from fracture surface.

Equatorial Scintillation Predictions from C/NOFS Planar Langmuir Probe Electron Density Fluctuation Data

DTIC Science & Technology

2014-09-05

Brasil Tel: 55 21 3527 1682; Fax: 55 21 3527 1154 E-mail: epoc@cetuc.puc-rio.br FINAL PERFORMANCE REPORT Equatorial scintillation...Costa Centro de Estudos em Telecomunicações/CETUC Rua Marquês de São Vicente, 225 – Gávea 22451-900 Rio de Janeiro - RJ – Brasil Tel: 55 21 3527...Emanoel Costa Centro de Estudos em Telecomunicações/CETUC Rua Marquês de São Vicente, 225 – Gávea 22451-900 Rio de Janeiro - RJ – Brasil Tel: 55

Electron microscopy methods in studies of cultural heritage sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vasiliev, A. L., E-mail: a.vasiliev56@gmail.com; Kovalchuk, M. V.; Yatsishina, E. B.

The history of the development and application of scanning electron microscopy (SEM), transmission electron microscopy (TEM), and energy-dispersive X-ray microanalysis (EDXMA) in studies of cultural heritage sites is considered. In fact, investigations based on these methods began when electron microscopes became a commercial product. Currently, these methods, being developed and improved, help solve many historical enigmas. To date, electron microscopy combined with microanalysis makes it possible to investigate any object, from parchment and wooden articles to pigments, tools, and objects of art. Studies by these methods have revealed that some articles were made by ancient masters using ancient “nanotechnologies”; hence,more » their comprehensive analysis calls for the latest achievements in the corresponding instrumental methods and sample preparation techniques.« less

Electron microscopy methods in studies of cultural heritage sites

NASA Astrophysics Data System (ADS)

Vasiliev, A. L.; Kovalchuk, M. V.; Yatsishina, E. B.

2016-11-01

The history of the development and application of scanning electron microscopy (SEM), transmission electron microscopy (TEM), and energy-dispersive X-ray microanalysis (EDXMA) in studies of cultural heritage sites is considered. In fact, investigations based on these methods began when electron microscopes became a commercial product. Currently, these methods, being developed and improved, help solve many historical enigmas. To date, electron microscopy combined with microanalysis makes it possible to investigate any object, from parchment and wooden articles to pigments, tools, and objects of art. Studies by these methods have revealed that some articles were made by ancient masters using ancient "nanotechnologies"; hence, their comprehensive analysis calls for the latest achievements in the corresponding instrumental methods and sample preparation techniques.

Comparison of dislocation content measured with transmission electron microscopy and micro-Laue diffraction based streak analysis

DOE PAGES

Zhang, C.; Balachandran, S.; Eisenlohr, P.; ...

2017-10-04

The subsurface dislocation content in a Ti-5Al-2.5Sn (wt%) uniaxial tension sample deformed at ambient temperature was characterized by peak streak analysis of micro-Laue diffraction patterns collected non-destructively by differential aperture X-raymicroscopy, and with focused ion beam transmission electron microscopy of material in the same volume. This comparison reveals that micro-Laue diffraction streak analysis based on an edge dislocation assumption can accurately identify the dominant dislocation slip system history (Burgers vector and plane observed by TEM), despite the fact that dislocations have predominantly screw character. As a result, other dislocations identified by TEM were not convincingly discernible from the peak streakmore » analysis.« less

TLM-Tracker: software for cell segmentation, tracking and lineage analysis in time-lapse microscopy movies.

PubMed

Klein, Johannes; Leupold, Stefan; Biegler, Ilona; Biedendieck, Rebekka; Münch, Richard; Jahn, Dieter

2012-09-01

Time-lapse imaging in combination with fluorescence microscopy techniques enable the investigation of gene regulatory circuits and uncovered phenomena like culture heterogeneity. In this context, computational image processing for the analysis of single cell behaviour plays an increasing role in systems biology and mathematical modelling approaches. Consequently, we developed a software package with graphical user interface for the analysis of single bacterial cell behaviour. A new software called TLM-Tracker allows for the flexible and user-friendly interpretation for the segmentation, tracking and lineage analysis of microbial cells in time-lapse movies. The software package, including manual, tutorial video and examples, is available as Matlab code or executable binaries at http://www.tlmtracker.tu-bs.de.

Comparison of dislocation content measured with transmission electron microscopy and micro-Laue diffraction based streak analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, C.; Balachandran, S.; Eisenlohr, P.

The subsurface dislocation content in a Ti-5Al-2.5Sn (wt%) uniaxial tension sample deformed at ambient temperature was characterized by peak streak analysis of micro-Laue diffraction patterns collected non-destructively by differential aperture X-raymicroscopy, and with focused ion beam transmission electron microscopy of material in the same volume. This comparison reveals that micro-Laue diffraction streak analysis based on an edge dislocation assumption can accurately identify the dominant dislocation slip system history (Burgers vector and plane observed by TEM), despite the fact that dislocations have predominantly screw character. As a result, other dislocations identified by TEM were not convincingly discernible from the peak streakmore » analysis.« less

Detecting Malaria Hotspots: A Comparison of Rapid Diagnostic Test, Microscopy, and Polymerase Chain Reaction

PubMed Central

Mogeni, Polycarp; Williams, Thomas N; Omedo, Irene; Kimani, Domtila; Ngoi, Joyce M; Mwacharo, Jedida; Morter, Richard; Nyundo, Christopher; Wambua, Juliana; Nyangweso, George; Kapulu, Melissa; Fegan, Gregory; Bejon, Philip

2017-01-01

Abstract Background Malaria control strategies need to respond to geographical hotspots of transmission. Detection of hotspots depends on the sensitivity of the diagnostic tool used. Methods We conducted cross-sectional surveys in 3 sites within Kilifi County, Kenya, that had variable transmission intensities. Rapid diagnostic test (RDT), microscopy, and polymerase chain reaction (PCR) were used to detect asymptomatic parasitemia, and hotspots were detected using the spatial scan statistic. Results Eight thousand five hundred eighty-one study participants were surveyed in 3 sites. There were statistically significant malaria hotspots by RDT, microscopy, and PCR for all sites except by microscopy in 1 low transmission site. Pooled data analysis of hotspots by PCR overlapped with hotspots by microscopy at a moderate setting but not at 2 lower transmission settings. However, variations in degree of overlap were noted when data were analyzed by year. Hotspots by RDT were predictive of PCR/microscopy at the moderate setting, but not at the 2 low transmission settings. We observed long-term stability of hotspots by PCR and microscopy but not RDT. Conclusion Malaria control programs may consider PCR testing to guide asymptomatic malaria hotspot detection once the prevalence of infection falls. PMID:28973672

Advanced techniques for in situ analysis of the biofilm matrix (structure, composition, dynamics) by means of laser scanning microscopy.

PubMed

Neu, Thomas R; Lawrence, John R

2014-01-01

The extracellular constituents in bioaggregates and biofilms can be imaged four dimensionally by using laser scanning microscopy. In this protocol we provide guidance on how to examine the various extracellular compartments in between microbial cells and communities associated with interfaces. The current options for fluorescence staining of matrix compounds and extracellular microhabitats are presented. Furthermore, practical aspects are discussed and useful notes are added. The chapter ends with a brief introduction to other approaches for EPS analysis and an outlook for future needs.

Optical contrast and refractive index of natural van der Waals heterostructure nanosheets of franckeite

PubMed Central

Gant, Patricia; Ghasemi, Foad; Maeso, David; Munuera, Carmen; López-Elvira, Elena; Frisenda, Riccardo; De Lara, David Pérez; Rubio-Bollinger, Gabino; Garcia-Hernandez, Mar

2017-01-01

We study mechanically exfoliated nanosheets of franckeite by quantitative optical microscopy. The analysis of transmission-mode and epi-illumination-mode optical microscopy images provides a rapid method to estimate the thickness of the exfoliated flakes at first glance. A quantitative analysis of the optical contrast spectra by means of micro-reflectance allows one to determine the refractive index of franckeite over a broad range of the visible spectrum through a fit of the acquired spectra to a model based on the Fresnel law. PMID:29181292

Quantitative study of mammalian cells by scanning transmission soft X-ray microscopy

NASA Astrophysics Data System (ADS)

Shinohara, K.; Ohigashi, T.; Toné, S.; Kado, M.; Ito, A.

2017-06-01

Molecular distribution in mammalian cells was studied by soft X-ray scanning transmission microscopy with respect to the quantitative aspect of analysis. NEXAFS profiles at the C, N and O K-absorption edges were combined and used for the analysis. For the estimation of quantity for nucleic acids and proteins, NEXAFS profiles of DNA and bovine serum albumin (BSA) at the N K-absorption edge were applied assuming that those were their representatives. The method has a potential to explore the other molecular components than nucleic acids and proteins.

Cost-effectiveness of novel algorithms for rapid diagnosis of tuberculosis in HIV-infected individuals in Uganda.

PubMed

Shah, Maunank; Dowdy, David; Joloba, Moses; Ssengooba, Willy; Manabe, Yukari C; Ellner, Jerrold; Dorman, Susan E

2013-11-28

Xpert MTB/RIF ('Xpert') and urinary lateral-flow lipoarabinomannan (LF-LAM) assays offer rapid tuberculosis (TB) diagnosis. This study evaluated the cost-effectiveness of novel diagnostic algorithms utilizing combinations of Xpert and LF-LAM for the detection of active TB among people living with HIV. Cost-effectiveness analysis using data from a comparative study of LF-LAM and Xpert, with a target population of HIV-infected individuals with signs/symptoms of TB in Uganda. A decision-analysis model compared multiple strategies for rapid TB diagnosis:sputum smear-microscopy; sputum Xpert; smear-microscopy combined with LF-LAM; and Xpert combined with LF-LAM. Primary outcomes were the costs and DALY's averted for each algorithm. Cost-effectiveness was represented using incremental cost-effectiveness ratios (ICER). Compared with an algorithm of Xpert testing alone, the combination of Xpert with LF-LAM was considered highly cost-effective (ICER $57/DALY-averted) at a willingness to pay threshold of Ugandan GDP per capita. Addition of urine LF-LAM testing to smear-microscopy was a less effective strategy than Xpert replacement of smear-microscopy, but was less costly and also considered highly cost-effective (ICER $33 per DALY-averted) compared with continued usage of smear-microscopy alone. Cost-effectiveness of the Xpert plus LF-LAM algorithm was most influenced by HIV/ART costs and life-expectancy of patients after TB treatment. The addition of urinary LF-LAM to TB diagnostic algorithms for HIV-infected individuals is highly cost-effective compared with usage of either sputum smear-microscopy or Xpert alone.

Enhanced polarizing microscopy as a new tool in aneuploidy research in oocytes.

PubMed

Shen, Ying; Betzendahl, Ilse; Tinneberg, Hans-Rudolf; Eichenlaub-Ritter, Ursula

2008-03-12

Chromosomal non-disjunction in female meiosis gives rise to reduced fertility and trisomy in humans. Human oocytes, especially from aged women, appear especially susceptible to non-disjunction. The oocyte spindle is crucial for high fidelity of chromosome segregation at meiotic divisions, and alterations in spindle morphology are therefore indicators of adverse conditions during oocyte development that may result in meiotic aneuploidy. In the past, oocytes had to be fixed for spindle analysis, precluding direct non-invasive identification of aneugens and adverse maturation conditions that affect spindle integrity and chromosome behaviour. Aneuploidy research for detection of spindle aberrations was therefore mainly focused on in vivo or in vitro exposed, fixed animal oocytes or cytogenetic analysis of spread oocytes. Orientation independent enhanced polarizing microscopy with nearly circularly polarized light and electronically controlled liquid crystal compensator optics is a new tool to study spindle morphology non-invasively in vivo for qualitative as well as quantitative analysis. Image generation by polarization microscopy depends on the intrinsic optical properties of the spindle with its paracrystalline microtubule lattice. When polarized light passes through such a lattice it induces a splitting of the beam and shift in the plane of vibration and retardation of light (termed birefringence and retardance). Studies of animal oocytes and follicle-cell denuded human oocytes fertilized by intracytoplasmic sperm injection for assisted conception have demonstrated the safety and efficacy of enhanced polarization microscopy. The method can be employed in aneuploidy research for non-invasive dose-response studies to detect spindle aberrations, for instance, in combination with cytogenetic analysis. Due to the non-invasive nature of the technique it may be employed in routine analysis of human oocytes to assess risks by lifestyle factors, and occupational and adverse environmental exposures.

Characterizing individual particles on tree leaves using computer automated scanning electron microscopy

Treesearch

D. L. Johnson; D. J. Nowak; V. A. Jouraeva

1999-01-01

Leaves from twenty-three deciduous tree species and five conifer species were collected within a limited geographic range (1 km radius) and evaluated for possible application of scanning electron microscopy and X-ray microanalysis techniques of individual particle analysis (IPA). The goal was to identify tree species with leaves suitable for the automated...

Observing Tin-Lead Alloys by Scanning Electron Microscopy: A Physical Chemistry Experiment Investigating Macro-Level Behaviors and Micro-Level Structures

ERIC Educational Resources Information Center

Wang, Yue; Xu, Xinhua; Wu, Meifen; Hu, Huikang; Wang, Xiaogang

2015-01-01

Scanning electron microscopy (SEM) was introduced into undergraduate physical chemistry laboratory curriculum to help students observe the phase composition and morphology characteristics of tin-lead alloys and thus further their understanding of binary alloy phase diagrams. The students were captivated by this visual analysis method, which…

SEM-EDX analysis of an unknown "known" white powder found in a shipping container from Peru

NASA Astrophysics Data System (ADS)

Albright, Douglas C.

2009-05-01

In 2008, an unknown white powder was discovered spilled inside of a shipping container of whole kernel corn during an inspection by federal inspectors in the port of Baltimore, Maryland. The container was detained and quarantined while a sample of the powder was collected and sent to a federal laboratory where it was screened using chromatography for the presence of specific poisons and pesticides with negative results. Samples of the corn kernels and the white powder were forwarded to the Food and Drug Administration, Forensic Chemistry Center for further analysis. Stereoscopic Light Microscopy (SLM), Scanning Electron Microscopy/Energy Dispersive X-ray Spectrometry (SEM/EDX), and Polarized Light Microscopy/Infrared Spectroscopy (PLM-IR) were used in the analysis of the kernels and the unknown powder. Based on the unique particle analysis by SLM and SEM as well as the detection of the presence of aluminum and phosphorous by EDX, the unknown was determined to be consistent with reacted aluminum phosphide (AlP). While commonly known in the agricultural industry, aluminum phosphide is relatively unknown in the forensic community. A history of the use and acute toxicity of this compound along with some very unique SEM/EDX analysis characteristics of aluminum phosphide will be discussed.

Mathematical imaging methods for mitosis analysis in live-cell phase contrast microscopy.

PubMed

Grah, Joana Sarah; Harrington, Jennifer Alison; Koh, Siang Boon; Pike, Jeremy Andrew; Schreiner, Alexander; Burger, Martin; Schönlieb, Carola-Bibiane; Reichelt, Stefanie

2017-02-15

In this paper we propose a workflow to detect and track mitotic cells in time-lapse microscopy image sequences. In order to avoid the requirement for cell lines expressing fluorescent markers and the associated phototoxicity, phase contrast microscopy is often preferred over fluorescence microscopy in live-cell imaging. However, common specific image characteristics complicate image processing and impede use of standard methods. Nevertheless, automated analysis is desirable due to manual analysis being subjective, biased and extremely time-consuming for large data sets. Here, we present the following workflow based on mathematical imaging methods. In the first step, mitosis detection is performed by means of the circular Hough transform. The obtained circular contour subsequently serves as an initialisation for the tracking algorithm based on variational methods. It is sub-divided into two parts: in order to determine the beginning of the whole mitosis cycle, a backwards tracking procedure is performed. After that, the cell is tracked forwards in time until the end of mitosis. As a result, the average of mitosis duration and ratios of different cell fates (cell death, no division, division into two or more daughter cells) can be measured and statistics on cell morphologies can be obtained. All of the tools are featured in the user-friendly MATLAB®Graphical User Interface MitosisAnalyser. Copyright © 2017. Published by Elsevier Inc.

Dimensional metrology of lab-on-a-chip internal structures: a comparison of optical coherence tomography with confocal fluorescence microscopy.

PubMed

Reyes, D R; Halter, M; Hwang, J

2015-07-01

The characterization of internal structures in a polymeric microfluidic device, especially of a final product, will require a different set of optical metrology tools than those traditionally used for microelectronic devices. We demonstrate that optical coherence tomography (OCT) imaging is a promising technique to characterize the internal structures of poly(methyl methacrylate) devices where the subsurface structures often cannot be imaged by conventional wide field optical microscopy. The structural details of channels in the devices were imaged with OCT and analyzed with an in-house written ImageJ macro in an effort to identify the structural details of the channel. The dimensional values obtained with OCT were compared with laser-scanning confocal microscopy images of channels filled with a fluorophore solution. Attempts were also made using confocal reflectance and interferometry microscopy to measure the channel dimensions, but artefacts present in the images precluded quantitative analysis. OCT provided the most accurate estimates for the channel height based on an analysis of optical micrographs obtained after destructively slicing the channel with a microtome. OCT may be a promising technique for the future of three-dimensional metrology of critical internal structures in lab-on-a-chip devices because scans can be performed rapidly and noninvasively prior to their use. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Survival and Intra-Nuclear Trafficking of Burkholderia pseudomallei: Strategies of Evasion from Immune Surveillance?

PubMed

Vadivelu, Jamuna; Vellasamy, Kumutha Malar; Thimma, Jaikumar; Mariappan, Vanitha; Kang, Wen-Tyng; Choh, Leang-Chung; Shankar, Esaki M; Wong, Kum Thong

2017-01-01

During infection, successful bacterial clearance is achieved via the host immune system acting in conjunction with appropriate antibiotic therapy. However, it still remains a tip of the iceberg as to where persistent pathogens namely, Burkholderia pseudomallei (B. pseudomallei) reside/hide to escape from host immune sensors and antimicrobial pressure. We used transmission electron microscopy (TEM) to investigate post-mortem tissue sections of patients with clinical melioidosis to identify the localisation of a recently identified gut microbiome, B. pseudomallei within host cells. The intranuclear presence of B. pseudomallei was confirmed using transmission electron microscopy (TEM) of experimentally infected guinea pig spleen tissues and Live Z-stack, and ImageJ analysis of fluorescence microscopy analysis of in vitro infection of A549 human lung epithelial cells. TEM investigations revealed intranuclear localization of B. pseudomallei in cells of infected human lung and guinea pig spleen tissues. We also found that B. pseudomallei induced actin polymerization following infection of A549 human lung epithelial cells. Infected A549 lung epithelial cells using 3D-Laser scanning confocal microscopy (LSCM) and immunofluorescence microscopy confirmed the intranuclear localization of B. pseudomallei. B. pseudomallei was found within the nuclear compartment of host cells. The nucleus may play a role as an occult or transient niche for persistence of intracellular pathogens, potentially leading to recurrrent episodes or recrudescence of infection.

Survival and Intra-Nuclear Trafficking of Burkholderia pseudomallei: Strategies of Evasion from Immune Surveillance?

PubMed Central

Vadivelu, Jamuna; Vellasamy, Kumutha Malar; Thimma, Jaikumar; Mariappan, Vanitha; Kang, Wen-Tyng; Choh, Leang-Chung; Wong, Kum Thong

2017-01-01

Background During infection, successful bacterial clearance is achieved via the host immune system acting in conjunction with appropriate antibiotic therapy. However, it still remains a tip of the iceberg as to where persistent pathogens namely, Burkholderia pseudomallei (B. pseudomallei) reside/hide to escape from host immune sensors and antimicrobial pressure. Methods We used transmission electron microscopy (TEM) to investigate post-mortem tissue sections of patients with clinical melioidosis to identify the localisation of a recently identified gut microbiome, B. pseudomallei within host cells. The intranuclear presence of B. pseudomallei was confirmed using transmission electron microscopy (TEM) of experimentally infected guinea pig spleen tissues and Live Z-stack, and ImageJ analysis of fluorescence microscopy analysis of in vitro infection of A549 human lung epithelial cells. Results TEM investigations revealed intranuclear localization of B. pseudomallei in cells of infected human lung and guinea pig spleen tissues. We also found that B. pseudomallei induced actin polymerization following infection of A549 human lung epithelial cells. Infected A549 lung epithelial cells using 3D-Laser scanning confocal microscopy (LSCM) and immunofluorescence microscopy confirmed the intranuclear localization of B. pseudomallei. Conclusion B. pseudomallei was found within the nuclear compartment of host cells. The nucleus may play a role as an occult or transient niche for persistence of intracellular pathogens, potentially leading to recurrrent episodes or recrudescence of infection. PMID:28045926

Inhomogeneity Based Characterization of Distribution Patterns on the Plasma Membrane

PubMed Central

Paparelli, Laura; Corthout, Nikky; Wakefield, Devin L.; Sannerud, Ragna; Jovanovic-Talisman, Tijana; Annaert, Wim; Munck, Sebastian

2016-01-01

Cell surface protein and lipid molecules are organized in various patterns: randomly, along gradients, or clustered when segregated into discrete micro- and nano-domains. Their distribution is tightly coupled to events such as polarization, endocytosis, and intracellular signaling, but challenging to quantify using traditional techniques. Here we present a novel approach to quantify the distribution of plasma membrane proteins and lipids. This approach describes spatial patterns in degrees of inhomogeneity and incorporates an intensity-based correction to analyze images with a wide range of resolutions; we have termed it Quantitative Analysis of the Spatial distributions in Images using Mosaic segmentation and Dual parameter Optimization in Histograms (QuASIMoDOH). We tested its applicability using simulated microscopy images and images acquired by widefield microscopy, total internal reflection microscopy, structured illumination microscopy, and photoactivated localization microscopy. We validated QuASIMoDOH, successfully quantifying the distribution of protein and lipid molecules detected with several labeling techniques, in different cell model systems. We also used this method to characterize the reorganization of cell surface lipids in response to disrupted endosomal trafficking and to detect dynamic changes in the global and local organization of epidermal growth factor receptors across the cell surface. Our findings demonstrate that QuASIMoDOH can be used to assess protein and lipid patterns, quantifying distribution changes and spatial reorganization at the cell surface. An ImageJ/Fiji plugin of this analysis tool is provided. PMID:27603951

Cement paste surface roughness analysis using coherence scanning interferometry and confocal microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Apedo, K.L., E-mail: apedo@unistra.fr; Munzer, C.; He, H.

2015-02-15

Scanning electron microscopy and scanning probe microscopy have been used for several decades to better understand the microstructure of cementitious materials. Very limited work has been performed to date to study the roughness of cementitious materials by optical microscopy such as coherence scanning interferometry (CSI) and chromatic confocal sensing (CCS). The objective of this paper is to better understand how CSI can be used as a tool to analyze surface roughness and topography of cement pastes. Observations from a series of images acquired using this technique on both polished and unpolished samples are described. The results from CSI are comparedmore » with those from a STIL confocal microscopy technique (SCM). Comparison between both optical techniques demonstrates the ability of CSI to measure both polished and unpolished cement pastes. - Highlights: • Coherence scanning interferometry (CSI) was used to analyze cement paste surfaces. • The results from the CSI were compared with those from a confocal microscopy. • 3D roughness parameters were obtained using the window resizing method. • Polished and unpolished cement pastes were studied.« less

Polychromatic flow cytometry is more sensitive than microscopy in detecting small monoclonal plasma cell populations.

PubMed

Tran, Daniel N; Smith, Sandy A B C; Brown, David A; Parker, Andrew J C; Joseph, Joanne E; Armstrong, Nicola; Sewell, William A

2017-03-01

There is an emerging role for flow cytometry (FC) in the assessment of small populations of plasma cells (PC). However, FC's utility has been questioned due to consistent underestimation of the percentage of PC compared to microscopy. A retrospective study was performed on bone marrow samples analysed by 8-colour FC. Plasma cell populations were classified as polyclonal or monoclonal based on FC analysis. FC findings were compared with microscopy of aspirates, histology and immunohistochemistry of trephine biopsies, and immunofixation (IFX) of serum and/or urine. FC underestimated PC compared to aspirate and trephine microscopy. The 10% diagnostic cutoff for MM on aspirate microscopy corresponded to a 3.5% cutoff on FC. Abnormal plasma cell morphology by aspirate microscopy and clonality by FC correlated in 229 of 294 cases (78%). However, in 50 cases, FC demonstrated a monoclonal population but microscopy reported no abnormality. In 15 cases, abnormalities were reported by microscopy but not by FC. Clonality assessment by trephine microscopy and FC agreed in 251/280 cases (90%), but all 29 discordant cases were monoclonal by FC and not monoclonal by microscopy. These cases had fewer PC and proportionally more polyclonal PC, and when IFX detected a paraprotein, it had the same light chain as in the PC determined by FC. FC was more sensitive in detecting monoclonal populations that were small or accompanied by polyclonal PC. This study supports the inclusion of FC in the evaluation of PC, especially in the assessment of small populations. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

Odontoma: retrospective study and confocal laser scanning microscope analysis of 52 cases.

PubMed

Crincoli, V; Scivetti, M; Di Bisceglie, M B; Lucchese, A; Favia, G

2007-01-01

The aim of this study was to perform a retrospective analysis of 52 cases of odontoma treated at the Department of Dentistry and Surgery, University of Bari, in the period 1971-2005. The odontogenic tumors were diagnosed as complex or compound odontoma following histological analysis and clinical radiological examination, and applying the 2005 WHO classification. The data analysis was conducted by considering the following factors: gender, age, site of the lesion, association with impacted teeth, aplasia, presence of supernumerary teeth as well as preoperative diagnosis by panoramic and periapical radiographs. Biopsy tissue samples were conventionally processed for histopathologic paraffin embedding and then were observed by optical microscopy and subsequently by confocal laser scanning microscopy (CLSM) in autofluorescence. Thirty specimens (57.6%) were from females and 22 (42.3%) were from males patients. The patients' age ranged from 5 to 75 years. Fifty-one percent of the specimens were excised from the mandible. In the maxilla, the most common location for odontomas was the anterior region. Most odontomas were associated with impacted teeth and only in one case there was an odontoma instead of a permanent tooth. Odontomas are considered hamartomatous malformations whose diagnosis is generally formulated by routinary radiographic examination. The CLSM analysis could help in diagnosis and histopathological analysis showing well-defined follicular area entrapped in hard tissues and pointing out ghost cells, otherwise not identifiable by traditional microscopy.

Theoretical, thermodynamic and electrochemical analysis of biotin drug as an impending corrosion inhibitor for mild steel in 15% hydrochloric acid

PubMed Central

Xu, Xihua; Sun, Zhipeng; Ansari, K. R.; Lin, Yuanhua

2017-01-01

The corrosion mitigation efficiency of biotin drug for mild steel in 15% hydrochloric acid was thoroughly investigated by weight loss and electrochemical methods. The surface morphology was studied by the contact angle, scanning electrochemical microscopy, atomic force microscopy and scanning electron microscopy methods. Quantum chemical calculation and Fukui analysis were done to correlate the experimental and theoretical data. The influence of the concentration of inhibitor, immersion time, temperature, activation energy, enthalpy and entropy has been reported. The mitigation efficiency of biotin obtained by all methods was in good correlation with each other. Polarization studies revealed that biotin acted as a mixed inhibitor. The adsorption of biotin was found to obey the Langmuir adsorption isotherm. Surface studies showed the hydrophobic nature of the steel with inhibitor and vindicated the formation of a film on the metal surface that reduced the corrosion rate. PMID:29308235

The Rényi divergence enables accurate and precise cluster analysis for localisation microscopy.

PubMed

Staszowska, Adela D; Fox-Roberts, Patrick; Hirvonen, Liisa M; Peddie, Christopher J; Collinson, Lucy M; Jones, Gareth E; Cox, Susan

2018-06-01

Clustering analysis is a key technique for quantitatively characterising structures in localisation microscopy images. To build up accurate information about biological structures, it is critical that the quantification is both accurate (close to the ground truth) and precise (has small scatter and is reproducible). Here we describe how the Rényi divergence can be used for cluster radius measurements in localisation microscopy data. We demonstrate that the Rényi divergence can operate with high levels of background and provides results which are more accurate than Ripley's functions, Voronoi tesselation or DBSCAN. Data supporting this research will be made accessible via a web link. Software codes developed for this work can be accessed via http://coxphysics.com/Renyi_divergence_software.zip. Implemented in C ++. Correspondence and requests for materials can be also addressed to the corresponding author. adela.staszowska@gmail.com or susan.cox@kcl.ac.uk. Supplementary data are available at Bioinformatics online.

SNOM and AFM microscopy techniques to study the effect of non-ionizing radiation on the morphological and biochemical properties of human keratinocytes cell line (HaCaT).

PubMed

Rieti, S; Manni, V; Lisi, A; Giuliani, L; Sacco, D; D'Emilia, E; Cricenti, A; Generosi, R; Luce, M; Grimaldi, S

2004-01-01

In this study we have employed atomic force microscopy (AFM) and scanning near-field optical microscopy (SNOM) techniques to study the effect of the interaction between human keratinocytes (HaCaT) and electromagnetic fields at low frequency. HaCaT cells were exposed to a sinusoidal magnetic field at a density of 50 Hz, 1 mT. AFM analysis revealed modification in shape and morphology in exposed cells with an increase in the areas of adhesion between cells. This latter finding was confirmed by SNOM indirect immunofluorescence analysis performed with a fluorescent antibody against the adhesion marker beta4 integrin, which revealed an increase of beta4 integrin segregation in the cell membrane of 50-Hz exposed cells, suggesting that a higher percentage of these cells shows a modified pattern of this adhesion marker.

The application of scanning electron microscopy to fractography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brooks, C.R.; McGill, B.L.

1994-10-01

Many failures involve fracture, and determination of the fracture process is a key factor in understanding the failure. This is frequently accomplished by characterizing the topography of the fracture surface. Scanning electron microscopy has a prominent role in fractography due to three features of the scanning electron microscope (SEM): high resolution, great depth of field, and the ability to obtain chemical information via analysis of the X-rays generated by the electrons. A qualitative treatment is presented of the interaction of electrons with a sample and the effect of the SEM operating parameters on image formation, quality, and X-ray analysis. Fractographsmore » are presented to illustrate these features of scanning electron microscopy and to illustrate the limitations and precautions in obtaining fractographs and x-ray analyses. The review is concluded with examples of fracture surface features of metallic, ceramic, and polymeric materials.« less

An historical account of the development and applications of the negative staining technique to the electron microscopy of viruses.

PubMed

Horne, R W; Wildy, P

1979-09-01

A brief historical account of the development and applications of the negative staining techniques to the study of the structure of viruses and their components as observed in the electron microscope is presented. Although the basic method of surrounding or embedding specimens in opaque dyes was used in light microscopy dating from about 1884, the equivalent preparative techniques applied to electron microscopy were comparatively recent. The combination of experiments on a sophisticated bacterial virus and the installation of a high resolution electron microscope in the Cavendish Laboratory, Cambridge, during 1954, subsequently led to the analysis of several important morphological features of animal, plant and bacterial viruses. The implications of the results from these early experiments on viruses and recent developments in negative staining methods for high resolution image analysis of electron micrographs are also discussed.

Analyzing Protein Clusters on the Plasma Membrane: Application of Spatial Statistical Analysis Methods on Super-Resolution Microscopy Images.

PubMed

Paparelli, Laura; Corthout, Nikky; Pavie, Benjamin; Annaert, Wim; Munck, Sebastian

2016-01-01

The spatial distribution of proteins within the cell affects their capability to interact with other molecules and directly influences cellular processes and signaling. At the plasma membrane, multiple factors drive protein compartmentalization into specialized functional domains, leading to the formation of clusters in which intermolecule interactions are facilitated. Therefore, quantifying protein distributions is a necessity for understanding their regulation and function. The recent advent of super-resolution microscopy has opened up the possibility of imaging protein distributions at the nanometer scale. In parallel, new spatial analysis methods have been developed to quantify distribution patterns in super-resolution images. In this chapter, we provide an overview of super-resolution microscopy and summarize the factors influencing protein arrangements on the plasma membrane. Finally, we highlight methods for analyzing clusterization of plasma membrane proteins, including examples of their applications.

Micro-XRF complemented by x-radiography and digital microscopy imaging for the study of hidden paintings

NASA Astrophysics Data System (ADS)

Gasanova, Svetlana; Hermon, Sorin

2017-07-01

The present study describes a novel approach to the study of hidden by integrating the non-invasive micro-X-Ray Fluorescence spectroscopy, X-radiography and digital microscopy. The case study analysed is a portrait of a male figure discovered under the painting of Ecce Homo, attributed to Titian's studio with an estimated date in the 1550s. The X-radiography images exposed the details of the underpainting, which appeared to be a nearly finished portrait of a standing man, overpainted by the current composition of Ecce Homo at a 180° angle. The microscopy observations of the upper painting's cracks and flaked areas enabled the study of the exposed underlayers in terms of their colour appearance and pigment particles. The subsequent pigment analysis was performed by micro-XRF. Since the described XRF analysis was performed not in scanner mode, the correct selection of the measurement spots for the micro analysis and separation between pigments of the lower and the upper painting was of paramount importance. The described approach for spot selection was based on the results of the preceding X-radiography and digital microscopy tests. The presence of lead white, vermilion, copper green and iron earth in the underlying portrait was confirmed by the multiple point XRF analysis of Pb, Hg, Cu, Fe and Mn lines. The described investigation method proved to be useful in the identification of the pigments of the underlying painting and consequently assisted in the tentative reconstruction of its colour palette. Moreover, the undertaken approach allowed discovering the potential of micro-XRF technique in the study of hidden compositions.

Structural Analysis of Enamel in Teeth from Head-and-Neck Cancer Patients Who Underwent Radiotherapy.

PubMed

Madrid, Cristhian C; de Pauli Paglioni, Mariana; Line, Sergio R; Vasconcelos, Karina G; Brandão, Thaís Bianca; Lopes, Marcio A; Santos-Silva, Alan Roger; De Goes, Mario Fernando

2017-01-01

To analyze macroscopic, microscopic, and ultrastructural aspects of enamel from head-and-neck cancer patients submitted to radiotherapy. Twenty sound extracted permanent molars were used and divided into 2 groups. The experimental group consisted of 10 molars from head-and-neck cancer patients submitted to radiotherapy with total doses that ranged from 50 to 70 Gy. Ten molars from patients who did not receive radiotherapy were matched with experimental-group samples by anatomic tooth group and comprised the control group. To perform a macroscopic analysis, standardized photos of different enamel faces were taken with a camera. Teeth were subjected to longitudinal cuts and hand polished to a final thickness of 0.1 mm. Enamel was analyzed under polarized light microscopy, and optical retardation values of birefringence were calculated in cervical, cusp, and occlusal pit areas. Subsequently, the same enamel areas were analyzed by scanning electron microscopy. Data from optical retardation values were statistically analyzed by 2-way ANOVA and Fisher's test (α < 0.05). No macroscopic differences were observed between the irradiated and control groups. Polarized light microscopy analysis revealed that cervical enamel exhibited darker areas characterized by discrete birefringence patterns compared to the control enamel. Optical retardation values were only significantly different in the cervical enamel of the irradiated and control groups (p < 0.0001). Scanning electron microscopy analysis revealed more evident interprismatic spaces in the cervical and outer cusp enamel of irradiated samples. Head-and-neck radiotherapy reduced optical retardation values of birefringence in cervical enamel, and the interprismatic spaces became more evident. © 2017 S. Karger AG, Basel.

Designing Image Analysis Pipelines in Light Microscopy: A Rational Approach.

PubMed

Arganda-Carreras, Ignacio; Andrey, Philippe

2017-01-01

With the progress of microscopy techniques and the rapidly growing amounts of acquired imaging data, there is an increased need for automated image processing and analysis solutions in biological studies. Each new application requires the design of a specific image analysis pipeline, by assembling a series of image processing operations. Many commercial or free bioimage analysis software are now available and several textbooks and reviews have presented the mathematical and computational fundamentals of image processing and analysis. Tens, if not hundreds, of algorithms and methods have been developed and integrated into image analysis software, resulting in a combinatorial explosion of possible image processing sequences. This paper presents a general guideline methodology to rationally address the design of image processing and analysis pipelines. The originality of the proposed approach is to follow an iterative, backwards procedure from the target objectives of analysis. The proposed goal-oriented strategy should help biologists to better apprehend image analysis in the context of their research and should allow them to efficiently interact with image processing specialists.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buck, E.C.; Cunnane, J.C.; Brown, N.R.

A combination of optical microscopy, scanning electron microscopy with backscattered electron detection (SEM/BSE), and analytical electron microscopy (AEM) is being used to determine the nature of uranium in soils from the Fernald Environmental Management Project. The information gained from these studies is being used to develop and test remediation technologies. Investigations using SEM have shown that uranium is contained within particles that are typically 1 to 100 {mu}m in diameter. Further analysis with AEM has shown that these uranium-rich regions are made up of discrete uranium-bearing phases. The distribution of these uranium phases was found to be inhomogeneous at themore » microscopic level.« less

Analysis of pigments in polychromes by use of laser induced breakdown spectroscopy and Raman microscopy

NASA Astrophysics Data System (ADS)

Castillejo, M.; Martín, M.; Silva, D.; Stratoudaki, T.; Anglos, D.; Burgio, L.; Clark, R. J. H.

2000-09-01

Two laser-based analytical techniques, Laser Induced Breakdown Spectroscopy (LIBS) and Raman microscopy, have been used for the identification of pigments on a polychrome from the Rococo period. Detailed spectral data are presented from analyses performed on a fragment of a gilded altarpiece from the church of Escatrón, Zaragoza, Spain. LIBS measurements yielded elemental analytical data which suggest the presence of certain pigments and, in addition, provide information on the stratigraphy of the paint layers. Identification of most pigments and of the materials used in the preparation layer was performed by Raman microscopy.

An observation of nanotwin lamellae in Cd 0.6Mn 0.4Te crystal by atomic force microscopy

NASA Astrophysics Data System (ADS)

George, M. A.; Azoulay, M.; Collins, W. E.; Burger, A.; Silberman, E.

1993-05-01

Atomic force microscopy (AFM) is used to examine the structure of freshly cleaved Cd 0.6Mn 0.4Te surfaces. The present report complements previous results obtained with X-ray diffraction and optical microscopy which showed the existence of microtwins. The AFM analysis was performed under ambient conditions and yielded nanometer scale resolution images of single twin lamellae that ranged between 20 and 100 nm in width. This is a first observation using AFM of such a substructure, which we interpret as evidence for the presence of nonotwins.

[Clinical pathology on the verge of virtual microscopy].

PubMed

Tolonen, Teemu; Näpänkangas, Juha; Isola, Jorma

2015-01-01

For more than 100 years, examinations of pathology specimens have relied on the use of the light microscope. The technological progress of the last few years is enabling the digitizing of histologic specimen slides and application of the virtual microscope in diagnostics. Virtual microscopy will facilitate consultation possibilities, and digital image analysis serves to enhance the level of diagnostics. Organizing and monitoring clinicopathological meetings will become easier. Digital archive of histologic specimens and the virtual microscopy network are expected to benefit training and research as well, particularly what applies to the Finnish biobank network which is currently being established.

Transmission electron microscopy analysis of skin lesions from sporotrichosis epidemic in Rio de Janeiro, Brazil.

PubMed

Ferreira, Cassio Porto; Oliveira de Almeida, Ana Cristina; Corte-Real, Suzana

2015-02-01

Transmission electron microscopy can yield useful information in a range of scientific fields; it is capable of imaging at a significantly higher resolution than light microscopes and has been a very useful tool in the identification of morphological changes of the dermis as well as assessment of changes in the extracellular matrix. Our aim is to characterize by electron microscopy the cellular profile of lesions caused by Sporothrix schenckii from the sporotrichosis epidemic in its zoonotic form that occurs in Rio de Janeiro, Brazil. © The American Society of Tropical Medicine and Hygiene.

FIMic: design for ultimate 3D-integral microscopy of in-vivo biological samples

PubMed Central

Scrofani, G.; Sola-Pikabea, J.; Llavador, A.; Sanchez-Ortiga, E.; Barreiro, J. C.; Saavedra, G.; Garcia-Sucerquia, J.; Martínez-Corral, M.

2017-01-01

In this work, Fourier integral microscope (FIMic), an ultimate design of 3D-integral microscopy, is presented. By placing a multiplexing microlens array at the aperture stop of the microscope objective of the host microscope, FIMic shows extended depth of field and enhanced lateral resolution in comparison with regular integral microscopy. As FIMic directly produces a set of orthographic views of the 3D-micrometer-sized sample, it is suitable for real-time imaging. Following regular integral-imaging reconstruction algorithms, a 2.75-fold enhanced depth of field and 2-time better spatial resolution in comparison with conventional integral microscopy is reported. Our claims are supported by theoretical analysis and experimental images of a resolution test target, cotton fibers, and in-vivo 3D-imaging of biological specimens. PMID:29359107

Study of electromechanical and mechanical properties of bacteria using force microscopy

NASA Astrophysics Data System (ADS)

Reukov, Vladimir; Thompson, Gary; Nikiforov, Maxim; Guo, Senli; Ovchinnikov, Oleg; Jesse, Stephen; Kalinin, Sergei; Vertegel, Alexey

2010-03-01

The application of scanning probe microscopy (SPM) to biological systems has evolved over the past decade into a multimodal and spectroscopic instrument that provides multiple information channels at each spatial pixel acquired. Recently, functional recognition imaging based on differing electromechanical properties between Gram negative and Gram positive bacteria was achieved using artificial neural network analysis of band excitation piezoresponse force microscopy (BEPFM) data. The immediate goal of this project was to study mechanical and electromechanical properties of bacterial systems physiologically-relevant solutions using Band-width Excitation Piezoresponce Force Microscopy (BE PFM) in combination with Force Mapping. Electromechanical imaging in physiological environments will improve the versatility of functional recognition imaging and open the way for application of the rapid BEPFM line mode method to other living cell systems.

Biological applications of phase-contrast electron microscopy.

PubMed

Nagayama, Kuniaki

2014-01-01

Here, I review the principles and applications of phase-contrast electron microscopy using phase plates. First, I develop the principle of phase contrast based on a minimal model of microscopy, introducing a double Fourier-transform process to mathematically formulate the image formation. Next, I explain four phase-contrast (PC) schemes, defocus PC, Zernike PC, Hilbert differential contrast, and schlieren optics, as image-filtering processes in the context of the minimal model, with particular emphases on the Zernike PC and corresponding Zernike phase plates. Finally, I review applications of Zernike PC cryo-electron microscopy to biological systems such as protein molecules, virus particles, and cells, including single-particle analysis to delineate three-dimensional (3D) structures of protein and virus particles and cryo-electron tomography to reconstruct 3D images of complex protein systems and cells.

Validation of an instrument for evaluating health care services to ostomized people.

PubMed

Moraes, Juliano Teixeira; Amaral, Carlos Faria Santos; Borges, Eline Lima; Ribeiro, Mauro Souza; Guimarães, Eliete Albano Azevedo

2016-12-08

to develop and validate an array of analysis and judgment for the evaluation of Health Care Services of people with stomas. cross-sectional study in 28 health facilities in the state of Minas Gerais. A descriptive analysis of the instrument and a study of its psychometric properties were performed. We used the Delphi technique for the validation of content and appearance. A psychometric analysis was carried out through the study of the reliability and validity of the measures obtained with the instrument. it was possible to construct an array analysis and judgment with 16 components (with scores from zero to five) grouped according to size and structure and process considered essential to evaluate the service. The results achieved in the reliability for structure and process, through the Cronbach alpha coefficient (α = 0.771 and α = 0.809, respectively), and the validity of content and construct demonstrated good internal consistency and satisfactory validity. An exploratory factor analysis indicated the item "main activity performed in the unit" as a limitation of the scale. the study provides a new tool for the evaluation of structure and process of Health Care Services of a Person with a stoma. desenvolver e validar uma matriz de análise e julgamento para a avaliação de Serviços de Atenção à Saúde da Pessoa Estomizada. estudo seccional em 28 unidades de saúde do estado de Minas Gerais. Foi realizada uma análise descritiva do instrumento e um estudo das suas propriedades psicométricas. Para a validação de conteúdo e de aparência foi utilizada a técnica Delphi. A análise psicométrica foi realizada por meio do estudo da confiabilidade e validade das medidas obtidas com o instrumento. foi possível construir uma matriz de análise e julgamento com 16 componentes (pontuados com escores de zero a cinco) agrupados de acordo com as dimensões estrutura e processo considerados fundamentais para a avaliação do serviço. Os resultados alcançados para a confiabilidade para estrutura e processo, por meio do Coeficiente alfa de Cronbach (α = 0,771 e α = 0,809 respectivamente), e validades de conteúdo e de construto, demonstraram boa consistência interna e satisfatória validade. A análise fatorial exploratória apontou o item "principal atividade realizada na unidade" como limitação da escala. o estudo disponibiliza nova ferramenta para a avaliação de estrutura e processo do Serviço de Atenção à Saúde da Pessoa Ostomizada. desarrollar y validar una matriz de análisis y juzgamiento para la evaluación de Servicios de Atención a la Salud de la Persona Ostomizada. estudio seccional en 28 unidades de salud del estado de Minas Gerais. Fue realizado un análisis descriptivo del instrumento y un estudio de sus propiedades psicométricas. Para la validación de contenido y de apariencia fue utilizada la técnica Delphi. El análisis psicométrico fue realizada por medio del estudio de la confiabilidad y validez de las medidas obtenidas con el instrumento. fue posible construir una matriz de análisis y juzgamiento con 16 componentes (evaluados con puntajes de cero a cinco) agrupados de acuerdo con las dimensiones estructura y proceso considerados fundamentales para la evaluación del servicio. Los resultados alcanzados para la confiabilidad para estructura y proceso, por medio del Coeficiente Alfa de Cronbach (α = 0,771 y α = 0,809 respectivamente), y validez de contenido y de constructo, demostraron buena consistencia interna y satisfactoria validez. El análisis factorial exploratória apuntó el ítem "principal actividad realizada en la unidad" como limitación de la escala. el estudio suministra una nueva herramienta para la evaluación de estructura y proceso del Servicio de Atención a la Salud de la Persona Ostomizada.

Surface-specific additive manufacturing test artefacts

NASA Astrophysics Data System (ADS)

Townsend, Andrew; Racasan, Radu; Blunt, Liam

2018-06-01

Many test artefact designs have been proposed for use with additive manufacturing (AM) systems. These test artefacts have primarily been designed for the evaluation of AM form and dimensional performance. A series of surface-specific measurement test artefacts designed for use in the verification of AM manufacturing processes are proposed here. Surface-specific test artefacts can be made more compact because they do not require the large dimensions needed for accurate dimensional and form measurements. The series of three test artefacts are designed to provide comprehensive information pertaining to the manufactured surface. Measurement possibilities include deviation analysis, surface texture parameter data generation, sub-surface analysis, layer step analysis and build resolution comparison. The test artefacts are designed to provide easy access for measurement using conventional surface measurement techniques, for example, focus variation microscopy, stylus profilometry, confocal microscopy and scanning electron microscopy. Additionally, the test artefacts may be simply visually inspected as a comparative tool, giving a fast indication of process variation between builds. The three test artefacts are small enough to be included in every build and include built-in manufacturing traceability information, making them a convenient physical record of the build.

Micrometer-scale particle sizing by laser diffraction: critical impact of the imaginary component of refractive index.

PubMed

Beekman, Alice; Shan, Daxian; Ali, Alana; Dai, Weiguo; Ward-Smith, Stephen; Goldenberg, Merrill

2005-04-01

This study evaluated the effect of the imaginary component of the refractive index on laser diffraction particle size data for pharmaceutical samples. Excipient particles 1-5 microm in diameter (irregular morphology) were measured by laser diffraction. Optical parameters were obtained and verified based on comparison of calculated vs. actual particle volume fraction. Inappropriate imaginary components of the refractive index can lead to inaccurate results, including false peaks in the size distribution. For laser diffraction measurements, obtaining appropriate or "effective" imaginary components of the refractive index was not always straightforward. When the recommended criteria such as the concentration match and the fit of the scattering data gave similar results for very different calculated size distributions, a supplemental technique, microscopy with image analysis, was used to decide between the alternatives. Use of effective optical parameters produced a good match between laser diffraction data and microscopy/image analysis data. The imaginary component of the refractive index can have a major impact on particle size results calculated from laser diffraction data. When performed properly, laser diffraction and microscopy with image analysis can yield comparable results.

Analysis of rolling fracture of the conticasted and tandem rolled blanks of low alloyed aluminum

NASA Astrophysics Data System (ADS)

Li, Yong; Zeng, Lingping; Jiao Xie, Xian

2018-01-01

Optical microscopy, electron microscopy and energy spectrum were used to test the morphology of grains, as-cast microstructure and secondary phases in confiscated and tandem rolled planks of 8011 low alloying aluminum alloy. It can be concluded that the existence of inhomogeneous secondary FeSiAl phases lead to the fracture of planks during rolling.

Evanescent Waves in High Numerical Aperture Aplanatic Solid Immersion Microscopy: Effects of Forbidden Light on Subsurface Imaging (Open Access, Publisher’s Version)

DTIC Science & Technology

2014-03-24

of the aSIL microscopy for semiconductor failure analysis and is applicable to imaging in quantum optics [18], biophotonics [19] and metrology [20...is usually of interest, the model can be adapted to applications in fields such as quantum optics and biophotonics for which the non-resonant

Determination of the refractive index of dehydrated cells by means of digital holographic microscopy

NASA Astrophysics Data System (ADS)

Belashov, A. V.; Zhikhoreva, A. A.; Bespalov, V. G.; Vasyutinskii, O. S.; Zhilinskaya, N. T.; Novik, V. I.; Semenova, I. V.

2017-10-01

Spatial distributions of the integral refractive index in dehydrated cells of human oral cavity epithelium are obtained by means of digital holographic microscopy, and mean refractive index of the cells is determined. The statistical analysis of the data obtained is carried out, and absolute errors of the method are estimated for different experimental conditions.

Advanced chemical imaging and comparison of human and porcine hair follicles for drug delivery by confocal Raman microscopy.

PubMed

Franzen, Lutz; Mathes, Christiane; Hansen, Steffi; Windbergs, Maike

2013-06-01

Hair follicles have recently gained a lot of interest for dermal drug delivery. They provide facilitated penetration into the skin and a high potential to serve as a drug depot. In this area of research, excised pig ear is a widely accepted in vitro model to evaluate penetration of drug delivery into hair follicles. However, a comparison of human and porcine follicles in terms of chemical composition has not been performed so far. In this study, we applied confocal Raman microscopy as a chemically selective imaging technique to compare human and porcine follicle composition and to visualize component distribution within follicle cross-sections. Based on the evaluation of human and porcine Raman spectra optical similarity for both species was successfully confirmed. Furthermore, cyanoacrylate skin surface biopsies, which are generally used to determine the extent of follicular penetration, were imaged by a novel complementary analytical approach combining confocal Raman microscopy and optical profilometry. This all-encompassing analysis allows investigation of intactness and component distribution of the excised hair bulb in three dimensions. Confocal Raman microscopy shows a high potential as a noninvasive and chemically selective technique for the analysis of trans-follicular drug delivery.

Advanced chemical imaging and comparison of human and porcine hair follicles for drug delivery by confocal Raman microscopy

NASA Astrophysics Data System (ADS)

Franzen, Lutz; Mathes, Christiane; Hansen, Steffi; Windbergs, Maike

2013-06-01

Hair follicles have recently gained a lot of interest for dermal drug delivery. They provide facilitated penetration into the skin and a high potential to serve as a drug depot. In this area of research, excised pig ear is a widely accepted in vitro model to evaluate penetration of drug delivery into hair follicles. However, a comparison of human and porcine follicles in terms of chemical composition has not been performed so far. In this study, we applied confocal Raman microscopy as a chemically selective imaging technique to compare human and porcine follicle composition and to visualize component distribution within follicle cross-sections. Based on the evaluation of human and porcine Raman spectra optical similarity for both species was successfully confirmed. Furthermore, cyanoacrylate skin surface biopsies, which are generally used to determine the extent of follicular penetration, were imaged by a novel complementary analytical approach combining confocal Raman microscopy and optical profilometry. This all-encompassing analysis allows investigation of intactness and component distribution of the excised hair bulb in three dimensions. Confocal Raman microscopy shows a high potential as a noninvasive and chemically selective technique for the analysis of trans-follicular drug delivery.

Background fluorescence estimation and vesicle segmentation in live cell imaging with conditional random fields.

PubMed

Pécot, Thierry; Bouthemy, Patrick; Boulanger, Jérôme; Chessel, Anatole; Bardin, Sabine; Salamero, Jean; Kervrann, Charles

2015-02-01

Image analysis applied to fluorescence live cell microscopy has become a key tool in molecular biology since it enables to characterize biological processes in space and time at the subcellular level. In fluorescence microscopy imaging, the moving tagged structures of interest, such as vesicles, appear as bright spots over a static or nonstatic background. In this paper, we consider the problem of vesicle segmentation and time-varying background estimation at the cellular scale. The main idea is to formulate the joint segmentation-estimation problem in the general conditional random field framework. Furthermore, segmentation of vesicles and background estimation are alternatively performed by energy minimization using a min cut-max flow algorithm. The proposed approach relies on a detection measure computed from intensity contrasts between neighboring blocks in fluorescence microscopy images. This approach permits analysis of either 2D + time or 3D + time data. We demonstrate the performance of the so-called C-CRAFT through an experimental comparison with the state-of-the-art methods in fluorescence video-microscopy. We also use this method to characterize the spatial and temporal distribution of Rab6 transport carriers at the cell periphery for two different specific adhesion geometries.

Form follows function: ultrastructure of different morphotypes of Physarum polycephalum

NASA Astrophysics Data System (ADS)

Oettmeier, Christina; Lee, Jonghyun; Döbereiner, Hans-Günther

2018-04-01

The multinucleate, unicellular slime mold Physarum polycephalum is a highly motile and morphologically diverse giant amoeba. Despite being brainless and lacking neurons, it exhibits ‘smart’ behavior. There is considerable interest in describing such traits and to investigate the underlying mechanochemical patterns which may hint at universal principles of behavior and decision-making. Furthermore, the slime mold’s mechanism of locomotion is unique. It resembles amoeboid movement, but differs from the locomotion of other amoebae in many ways, e.g. in their much larger size and lack of lobopodia. These two aspects, behavior and locomotion, are linked by the cytoskeleton and the overall morphology of P. polycephalum. In this paper, we present a structural analysis of different growth forms (micro-, meso- and macroplasmodia) by transmission electron microscopy (TEM), scanning electron microscopy (SEM), light microscopy, and fluorescence microscopy of F-actin. With these detailed investigations of cellular ultrastructure and morphology, we provide the basis for the analysis of, e.g. viscoelastic and rheological measurements. Our data also provide structural details for the many models that have been constructed for the understanding of locomotion. We conclude that morphological information is vital for the assessment and measurement of material properties.

The role of molecular genetic analysis in the diagnosis of primary ciliary dyskinesia.

PubMed

Kim, Raymond H; A Hall, David; Cutz, Ernest; Knowles, Michael R; Nelligan, Kathleen A; Nykamp, Keith; Zariwala, Maimoona A; Dell, Sharon D

2014-03-01

Primary ciliary dyskinesia (PCD) is an autosomal recessive genetic disorder of motile cilia. The diagnosis of PCD has previously relied on ciliary analysis with transmission electron microscopy or video microscopy. However, patients with PCD may have normal ultrastructural appearance, and ciliary analysis has limited accessibility. Alternatively, PCD can be diagnosed by demonstrating biallelic mutations in known PCD genes. Genetic testing is emerging as a diagnostic tool to complement ciliary analysis where interpretation and access may delay diagnosis. To determine the diagnostic yield of genetic testing of patients with a confirmed or suspected diagnosis of PCD in a multiethnic urban center. Twenty-eight individuals with confirmed PCD on transmission electron microscopy of ciliary ultrastructure and 24 individuals with a probable diagnosis of PCD based on a classical PCD phenotype and low nasal nitric oxide had molecular analysis of 12 genes associated with PCD. Of 49 subjects who underwent ciliary biopsy, 28 (57%) were diagnosed with PCD through an ultrastructural defect. Of the 52 individuals who underwent molecular genetic analysis, 22 (42%) individuals had two mutations in known PCD genes. Twenty-four previously unreported mutations in known PCD genes were observed. Combining both diagnostic modalities of biopsy and molecular genetics, the diagnostic yield increased to 69% compared with 57% based on biopsy alone. The diagnosis of PCD is challenging and has traditionally relied on ciliary biopsy, which is unreliable as the sole criterion for a definitive diagnosis. Molecular genetic analysis can be used as a complementary test to increase the diagnostic yield.

Correlative Light- and Electron Microscopy Using Quantum Dot Nanoparticles.

PubMed

Killingsworth, Murray C; Bobryshev, Yuri V

2016-08-07

A method is described whereby quantum dot (QD) nanoparticles can be used for correlative immunocytochemical studies of human pathology tissue using widefield fluorescence light microscopy and transmission electron microscopy (TEM). To demonstrate the protocol we have immunolabeled ultrathin epoxy sections of human somatostatinoma tumor using a primary antibody to somatostatin, followed by a biotinylated secondary antibody and visualization with streptavidin conjugated 585 nm cadmium-selenium (CdSe) quantum dots (QDs). The sections are mounted on a TEM specimen grid then placed on a glass slide for observation by widefield fluorescence light microscopy. Light microscopy reveals 585 nm QD labeling as bright orange fluorescence forming a granular pattern within the tumor cell cytoplasm. At low to mid-range magnification by light microscopy the labeling pattern can be easily recognized and the level of non-specific or background labeling assessed. This is a critical step for subsequent interpretation of the immunolabeling pattern by TEM and evaluation of the morphological context. The same section is then blotted dry and viewed by TEM. QD probes are seen to be attached to amorphous material contained in individual secretory granules. Images are acquired from the same region of interest (ROI) seen by light microscopy for correlative analysis. Corresponding images from each modality may then be blended to overlay fluorescence data on TEM ultrastructure of the corresponding region.

Real-time, non-invasive microscopic confirmation of clinical diagnosis of bullous pemphigoid using in vivo reflectance confocal microscopy.

PubMed

Ardigò, M; Agozzino, M; Amorosi, B; Moscarella, E; Cota, C; de Abreu, L; Berardesca, E

2014-05-01

Bullous pemphigoid is an autoimmune disease affecting prevalently the elder. In vivo reflectance confocal microscopy is a non-invasive technique for real-time imaging of the skin with cellular-level resolution. No previous data has been reported about confocal microscopy of bullous pemphigoid. Aim of this preliminary study is the evaluation of the potential of in vivo reflectance confocal microscopy for real-time, microscopical confirmation of clinical bullous pemphigoid diagnosis. A total of nine lesions from patients affected by pemphigoid underwent in vivo reflectance confocal microscopy before histological examination. In our preliminary study, confocal microscopy showed high grade of correspondence to histopathology. In particular, presence of sub-epidermal cleft and variable amount of oedema of the upper dermis associated with inflammatory cells infiltration were seen as prevalent confocal features in the bullous lesions considered. Differently, in urticarial lesions, no specific features could be appreciated at confocal analysis beside the presence of signs of spongiosis and perivascular inflammation. Confocal microscopy seems to be useful for in vivo, microscopical confirmation of the clinical suspect of bullous pemphigoid and for biopsy site selection in urticarial lesions to obtain a more significant specimen for histopathological examination. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Bending the Rules: Widefield Microscopy and the Abbe Limit of Resolution

PubMed Central

Verdaasdonk, Jolien S.; Stephens, Andrew D.; Haase, Julian; Bloom, Kerry

2014-01-01

One of the most fundamental concepts of microscopy is that of resolution–the ability to clearly distinguish two objects as separate. Recent advances such as structured illumination microscopy (SIM) and point localization techniques including photoactivated localization microscopy (PALM), and stochastic optical reconstruction microscopy (STORM) strive to overcome the inherent limits of resolution of the modern light microscope. These techniques, however, are not always feasible or optimal for live cell imaging. Thus, in this review, we explore three techniques for extracting high resolution data from images acquired on a widefield microscope–deconvolution, model convolution, and Gaussian fitting. Deconvolution is a powerful tool for restoring a blurred image using knowledge of the point spread function (PSF) describing the blurring of light by the microscope, although care must be taken to ensure accuracy of subsequent quantitative analysis. The process of model convolution also requires knowledge of the PSF to blur a simulated image which can then be compared to the experimentally acquired data to reach conclusions regarding its geometry and fluorophore distribution. Gaussian fitting is the basis for point localization microscopy, and can also be applied to tracking spot motion over time or measuring spot shape and size. All together, these three methods serve as powerful tools for high-resolution imaging using widefield microscopy. PMID:23893718

Detecting Malaria Hotspots: A Comparison of Rapid Diagnostic Test, Microscopy, and Polymerase Chain Reaction.

PubMed

Mogeni, Polycarp; Williams, Thomas N; Omedo, Irene; Kimani, Domtila; Ngoi, Joyce M; Mwacharo, Jedida; Morter, Richard; Nyundo, Christopher; Wambua, Juliana; Nyangweso, George; Kapulu, Melissa; Fegan, Gregory; Bejon, Philip

2017-11-27

Malaria control strategies need to respond to geographical hotspots of transmission. Detection of hotspots depends on the sensitivity of the diagnostic tool used. We conducted cross-sectional surveys in 3 sites within Kilifi County, Kenya, that had variable transmission intensities. Rapid diagnostic test (RDT), microscopy, and polymerase chain reaction (PCR) were used to detect asymptomatic parasitemia, and hotspots were detected using the spatial scan statistic. Eight thousand five hundred eighty-one study participants were surveyed in 3 sites. There were statistically significant malaria hotspots by RDT, microscopy, and PCR for all sites except by microscopy in 1 low transmission site. Pooled data analysis of hotspots by PCR overlapped with hotspots by microscopy at a moderate setting but not at 2 lower transmission settings. However, variations in degree of overlap were noted when data were analyzed by year. Hotspots by RDT were predictive of PCR/microscopy at the moderate setting, but not at the 2 low transmission settings. We observed long-term stability of hotspots by PCR and microscopy but not RDT. Malaria control programs may consider PCR testing to guide asymptomatic malaria hotspot detection once the prevalence of infection falls. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Laser scanning saturated structured illumination microscopy based on phase modulation

NASA Astrophysics Data System (ADS)

Huang, Yujia; Zhu, Dazhao; Jin, Luhong; Kuang, Cuifang; Xu, Yingke; Liu, Xu

2017-08-01

Wide-field saturated structured illumination microscopy has not been widely used due to the requirement of high laser power. We propose a novel method called laser scanning saturated structured illumination microscopy (LS-SSIM), which introduces high order of harmonics frequency and greatly reduces the required laser power for SSIM imaging. To accomplish that, an excitation PSF with two peaks is generated and scanned along different directions on the sample. Raw images are recorded cumulatively by a CCD detector and then reconstructed to form a high-resolution image with extended optical transfer function (OTF). Our theoretical analysis and simulation results show that LS-SSIM method reaches a resolution of 0.16 λ, equivalent to 2.7-fold resolution than conventional wide-field microscopy. In addition, LS-SSIM greatly improves the optical sectioning capability of conventional wide-field illumination system by diminishing our-of-focus light. Furthermore, this modality has the advantage of implementation in multi-photon microscopy with point scanning excitation to image samples in greater depths.

Identifying Nanoscale Structure-Function Relationships Using Multimodal Atomic Force Microscopy, Dimensionality Reduction, and Regression Techniques.

PubMed

Kong, Jessica; Giridharagopal, Rajiv; Harrison, Jeffrey S; Ginger, David S

2018-05-31

Correlating nanoscale chemical specificity with operational physics is a long-standing goal of functional scanning probe microscopy (SPM). We employ a data analytic approach combining multiple microscopy modes, using compositional information in infrared vibrational excitation maps acquired via photoinduced force microscopy (PiFM) with electrical information from conductive atomic force microscopy. We study a model polymer blend comprising insulating poly(methyl methacrylate) (PMMA) and semiconducting poly(3-hexylthiophene) (P3HT). We show that PiFM spectra are different from FTIR spectra, but can still be used to identify local composition. We use principal component analysis to extract statistically significant principal components and principal component regression to predict local current and identify local polymer composition. In doing so, we observe evidence of semiconducting P3HT within PMMA aggregates. These methods are generalizable to correlated SPM data and provide a meaningful technique for extracting complex compositional information that are impossible to measure from any one technique.

Towards native-state imaging in biological context in the electron microscope

PubMed Central

Weston, Anne E.; Armer, Hannah E. J.

2009-01-01

Modern cell biology is reliant on light and fluorescence microscopy for analysis of cells, tissues and protein localisation. However, these powerful techniques are ultimately limited in resolution by the wavelength of light. Electron microscopes offer much greater resolution due to the shorter effective wavelength of electrons, allowing direct imaging of sub-cellular architecture. The harsh environment of the electron microscope chamber and the properties of the electron beam have led to complex chemical and mechanical preparation techniques, which distance biological samples from their native state and complicate data interpretation. Here we describe recent advances in sample preparation and instrumentation, which push the boundaries of high-resolution imaging. Cryopreparation, cryoelectron microscopy and environmental scanning electron microscopy strive to image samples in near native state. Advances in correlative microscopy and markers enable high-resolution localisation of proteins. Innovation in microscope design has pushed the boundaries of resolution to atomic scale, whilst automatic acquisition of high-resolution electron microscopy data through large volumes is finally able to place ultrastructure in biological context. PMID:19916039

Q-controlled amplitude modulation atomic force microscopy in liquids: An analysis

NASA Astrophysics Data System (ADS)

Hölscher, H.; Schwarz, U. D.

2006-08-01

An analysis of amplitude modulation atomic force microscopy in liquids is presented with respect to the application of the Q-Control technique. The equation of motion is solved by numerical and analytic methods with and without Q-Control in the presence of a simple model interaction force adequate for many liquid environments. In addition, the authors give an explicit analytical formula for the tip-sample indentation showing that higher Q factors reduce the tip-sample force. It is found that Q-Control suppresses unwanted deformations of the sample surface, leading to the enhanced image quality reported in several experimental studies.

Preparation and Microcosmic Structural Analysis of Recording Coating on Inkjet Printing Media

PubMed Central

Jiang, Bo; Liu, Weiyan; Bai, Yongping; Huang, Yudong; Liu, Li; Han, Jianping

2011-01-01

Preparation of recording coating on inkjet printing (RC-IJP) media was proposed. The microstructure and roughness of RC-IJP was analyzed by scanning electron microscopy (SEM) and atomic force microscope (AFM). The surface infiltration process of RC-IJP was studied by a liquid infiltration instrument. The distribution of C, O and Si composites on recording coating surface is analyzed by energy dispersive spectrum (EDS). The transmission electron microscopy (TEM) analysis showed that the nanoscale silica could be dissolved uniformly in water. Finally, the print color is shown clearly by the preparative recording coating. PMID:21954368

Visual-servoing optical microscopy

DOEpatents

Callahan, Daniel E.; Parvin, Bahram

2009-06-09

The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time: quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

Visual-servoing optical microscopy

DOEpatents

Callahan, Daniel E [Martinez, CA; Parvin, Bahram [Mill Valley, CA

2011-05-24

The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time; quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

Visual-servoing optical microscopy

DOEpatents

Callahan, Daniel E; Parvin, Bahram

2013-10-01

The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time; quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

Analysis of Multilayer Devices for Superconducting Electronics by High-Resolution Scanning Transmission Electron Microscopy and Energy Dispersive Spectroscopy

DOE PAGES

Missert, Nancy; Kotula, Paul G.; Rye, Michael; ...

2017-02-15

We used a focused ion beam to obtain cross-sectional specimens from both magnetic multilayer and Nb/Al-AlOx/Nb Josephson junction devices for characterization by scanning transmission electron microscopy (STEM) and energy dispersive X-ray spectroscopy (EDX). An automated multivariate statistical analysis of the EDX spectral images produced chemically unique component images of individual layers within the multilayer structures. STEM imaging elucidated distinct variations in film morphology, interface quality, and/or etch artifacts that could be correlated to magnetic and/or electrical properties measured on the same devices.

Nomarski differential interference contrast microscopy for surface slope measurements: an examination of techniques

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hartman, J.S.; Gordon, R.L.; Lessor, D.L.

1981-08-01

Alternate measurement and data analysis procedures are discussed and compared for the application of reflective Nomarski differential interference contrast microscopy for the determination of surface slopes. The discussion includes the interpretation of a previously reported iterative procedure using the results of a detailed optical model and the presentation of a new procedure based on measured image intensity extrema. Surface slope determinations from these procedures are presented and compared with results from a previously reported curve fit analysis of image intensity data. The accuracy and advantages of the different procedures are discussed.

Doxorubicin-loaded Zein in situ gel for interstitial chemotherapy.

PubMed

Cao, Xiaoying; Geng, Jianning; Su, Suwen; Zhang, Linan; Xu, Qian; Zhang, Li; Xie, Yinghua; Wu, Shaomei; Sun, Yongjun; Gao, Zibin

2012-01-01

A novel drug delivery system of doxorubicin (DOX)-loaded Zein in situ gel for interstitial chemotherapy was investigated in this study. The possible mechanisms of drug release were described according to morphological analysis by optical microscopy and scanning electronic microscope (SEM). In vitro and in vivo anti-tumor activity studies showed that DOX-loaded Zein in situ gel was superior to DOX solution. Local pharmacokinetics in tumor tissue was studied by quantitative analysis with confocal laser scanning microscopy (CLSM) combined with microdialysis technology. A pharmacokinetics mathematical model of DOX-loaded Zein in situ gel in tumors was then built.

Surface diagnostics for scale analysis.

PubMed

Dunn, S; Impey, S; Kimpton, C; Parsons, S A; Doyle, J; Jefferson, B

2004-01-01

Stainless steel, polymethylmethacrylate and polytetrafluoroethylene coupons were analysed for surface topographical and adhesion force characteristics using tapping mode atomic force microscopy and force-distance microscopy techniques. The two polymer materials were surface modified by polishing with silicon carbide papers of known grade. The struvite scaling rate was determined for each coupon and related to the data gained from the surface analysis. The scaling rate correlated well with adhesion force measurements indicating that lower energy materials scale at a lower rate. The techniques outlined in the paper provide a method for the rapid screening of materials in potential scaling applications.

Translation, cultural adaptation and validation of Kidney Disease Loss Scale to the Brazilian context.

PubMed

Ottaviani, Ana Carolina; Orlandi, Fabiana de Souza

2016-01-01

Losses can be conceptualized as cognitive and affective responses to individual sorrows, characterized by brooding, yearning, disbelief and stunned feelings, being clinically significant in chronic diseases. The aim of the study was to translate, culturally adapt and validate the Kidney Disease Loss Scale into Portuguese. Validation study involving the steps recommended in the literature for healthcare instruments: initial translation, synthesis of translations, back translation, review by a committee of judges and pretest. The scale was translated and adapted to the Portuguese language, being quick and easy to application. The reliability and reproducibility showed satisfactory values. Factor analysis indicated a factor that explains 59.7% of the losses construct. The Kidney Disease Loss Scale was translated, adapted and validated for the Brazilian context, allowing future studies of losses and providing tools for the professionals working in dialysis centers for assistance to people with chronic kidney disease. As perdas podem ser conceituadas como respostas cognitivas e afetivas para tristezas individuais, caracterizadas pelo remoer, anseio, descrença e sentimentos atordoados, sendo clinicamente significativa em doenças crônicas. O objetivo do estudo foi traduzir, adaptar culturalmente e validar o Kidney Disease Loss Scale para a língua portuguesa. Estudo de validação envolveu as etapas preconizadas na literatura internacional para instrumentos da área de saúde: tradução inicial, síntese das traduções, retrotradução, revisão por um comitê de juízes, pré-teste e avaliação das propriedades psicométricas. A escala foi traduzida e adaptada para o idioma português, sendo de fácil e rápida aplicação. A confiabilidade e a reprodutibilidade apresentaram valores satisfatórios. A análise fatorial indicou um fator que explica 59,7% do constructo de perdas. A Escala de Perdas referente à Doença Renal foi traduzida, adaptada e validada para o contexto brasileiro, permitindo estudos futuros sobre perdas e instrumentalizando os profissionais atuantes em centros de diálise para assistência à pessoa com doença renal crônica.

Image correlation microscopy for uniform illumination.

PubMed

Gaborski, T R; Sealander, M N; Ehrenberg, M; Waugh, R E; McGrath, J L

2010-01-01

Image cross-correlation microscopy is a technique that quantifies the motion of fluorescent features in an image by measuring the temporal autocorrelation function decay in a time-lapse image sequence. Image cross-correlation microscopy has traditionally employed laser-scanning microscopes because the technique emerged as an extension of laser-based fluorescence correlation spectroscopy. In this work, we show that image correlation can also be used to measure fluorescence dynamics in uniform illumination or wide-field imaging systems and we call our new approach uniform illumination image correlation microscopy. Wide-field microscopy is not only a simpler, less expensive imaging modality, but it offers the capability of greater temporal resolution over laser-scanning systems. In traditional laser-scanning image cross-correlation microscopy, lateral mobility is calculated from the temporal de-correlation of an image, where the characteristic length is the illuminating laser beam width. In wide-field microscopy, the diffusion length is defined by the feature size using the spatial autocorrelation function. Correlation function decay in time occurs as an object diffuses from its original position. We show that theoretical and simulated comparisons between Gaussian and uniform features indicate the temporal autocorrelation function depends strongly on particle size and not particle shape. In this report, we establish the relationships between the spatial autocorrelation function feature size, temporal autocorrelation function characteristic time and the diffusion coefficient for uniform illumination image correlation microscopy using analytical, Monte Carlo and experimental validation with particle tracking algorithms. Additionally, we demonstrate uniform illumination image correlation microscopy analysis of adhesion molecule domain aggregation and diffusion on the surface of human neutrophils.

Non-linear imaging and characterization of atherosclerotic arterial tissue using combined two photon fluorescence, second-harmonic generation and CARS microscopy

NASA Astrophysics Data System (ADS)

Cicchi, Riccardo; Matthäus, Christian; Meyer, Tobias; Lattermann, Annika; Dietzek, Benjamin; Brehm, Bernhard R.; Popp, Jürgen; Pavone, Francesco S.

2014-02-01

Atherosclerosis is among the most widespread cardiovascular diseases and one of the leading cause of death in the Western World. Characterization of arterial tissue in atherosclerotic condition is extremely interesting from the diagnostic point of view. Routinely used diagnostic methods, such as histopathological examination, are limited to morphological analysis of the examined tissues, whereas an exhaustive characterization requires a morpho-functional approach. Multimodal non-linear microscopy has the potential to bridge this gap by providing morpho-functional information on the examined tissues in a label-free way. Here we employed multiple non-linear microscopy techniques, including CARS, TPF, and SHG to provide intrinsic optical contrast from various tissue components in both arterial wall and atherosclerotic plaques. CARS and TPF microscopy were used to respectively image lipid depositions within plaques and elastin in the arterial wall. Cholesterol deposition in the lumen and collagen in the arterial wall were selectively imaged by SHG microscopy and distinguished by forward-backward SHG ratio. Image pattern analysis allowed characterizing collagen organization in different tissue regions. Different values of fiber mean size, distribution and anisotropy are calculated for lumen and media prospectively allowing for automated classification of atherosclerotic lesions. The presented method represents a promising diagnostic tool for evaluating atherosclerotic tissue and has the potential to find a stable place in clinical setting as well as to be applied in vivo in the near future.

Next-generation endomyocardial biopsy: the potential of confocal and super-resolution microscopy.

PubMed

Crossman, David J; Ruygrok, Peter N; Hou, Yu Feng; Soeller, Christian

2015-03-01

Confocal laser scanning microscopy and super-resolution microscopy provide high-contrast and high-resolution fluorescent imaging, which has great potential to increase the diagnostic yield of endomyocardial biopsy (EMB). EMB is currently the gold standard for identification of cardiac allograft rejection, myocarditis, and infiltrative and storage diseases. However, standard analysis is dominated by low-contrast bright-field light and electron microscopy (EM); this lack of contrast makes quantification of pathological features difficult. For example, assessment of cardiac allograft rejection relies on subjective grading of H&E histology, which may lead to diagnostic variability between pathologists. This issue could be solved by utilising the high contrast provided by fluorescence methods such as confocal to quantitatively assess the degree of lymphocytic infiltrate. For infiltrative diseases such as amyloidosis, the nanometre resolution provided by EM can be diagnostic in identifying disease-causing fibrils. The recent advent of super-resolution imaging, particularly direct stochastic optical reconstruction microscopy (dSTORM), provides high-contrast imaging at resolution approaching that of EM. Moreover, dSTORM utilises conventional fluorescence dyes allowing for the same structures to be routinely imaged at the cellular scale and then at the nanoscale. The key benefit of these technologies is that the high contrast facilitates quantitative digital analysis and thereby provides a means to robustly assess critical pathological features. Ultimately, this technology has the ability to provide greater accuracy and precision to EMB assessment, which could result in better outcomes for patients.

Bleed-through correction for rendering and correlation analysis in multi-colour localization microscopy

PubMed Central

Kim, Dahan; Curthoys, Nikki M.; Parent, Matthew T.; Hess, Samuel T.

2015-01-01

Multi-colour localization microscopy has enabled sub-diffraction studies of colocalization between multiple biological species and quantification of their correlation at length scales previously inaccessible with conventional fluorescence microscopy. However, bleed-through, or misidentification of probe species, creates false colocalization and artificially increases certain types of correlation between two imaged species, affecting the reliability of information provided by colocalization and quantified correlation. Despite the potential risk of these artefacts of bleed-through, neither the effect of bleed-through on correlation nor methods of its correction in correlation analyses has been systematically studied at typical rates of bleed-through reported to affect multi-colour imaging. Here, we present a reliable method of bleed-through correction applicable to image rendering and correlation analysis of multi-colour localization microscopy. Application of our bleed-through correction shows our method accurately corrects the artificial increase in both types of correlations studied (Pearson coefficient and pair correlation), at all rates of bleed-through tested, in all types of correlations examined. In particular, anti-correlation could not be quantified without our bleed-through correction, even at rates of bleed-through as low as 2%. Demonstrated with dichroic-based multi-colour FPALM here, our presented method of bleed-through correction can be applied to all types of localization microscopy (PALM, STORM, dSTORM, GSDIM, etc.), including both simultaneous and sequential multi-colour modalities, provided the rate of bleed-through can be reliably determined. PMID:26185614

Bleed-through correction for rendering and correlation analysis in multi-colour localization microscopy.

PubMed

Kim, Dahan; Curthoys, Nikki M; Parent, Matthew T; Hess, Samuel T

2013-09-01

Multi-colour localization microscopy has enabled sub-diffraction studies of colocalization between multiple biological species and quantification of their correlation at length scales previously inaccessible with conventional fluorescence microscopy. However, bleed-through, or misidentification of probe species, creates false colocalization and artificially increases certain types of correlation between two imaged species, affecting the reliability of information provided by colocalization and quantified correlation. Despite the potential risk of these artefacts of bleed-through, neither the effect of bleed-through on correlation nor methods of its correction in correlation analyses has been systematically studied at typical rates of bleed-through reported to affect multi-colour imaging. Here, we present a reliable method of bleed-through correction applicable to image rendering and correlation analysis of multi-colour localization microscopy. Application of our bleed-through correction shows our method accurately corrects the artificial increase in both types of correlations studied (Pearson coefficient and pair correlation), at all rates of bleed-through tested, in all types of correlations examined. In particular, anti-correlation could not be quantified without our bleed-through correction, even at rates of bleed-through as low as 2%. Demonstrated with dichroic-based multi-colour FPALM here, our presented method of bleed-through correction can be applied to all types of localization microscopy (PALM, STORM, dSTORM, GSDIM, etc.), including both simultaneous and sequential multi-colour modalities, provided the rate of bleed-through can be reliably determined.

Identification and quantitive analysis of calcium phosphate microparticles in intestinal tissue by nuclear microscopy

NASA Astrophysics Data System (ADS)

Gomez-Morilla, Inmaculada; Thoree, Vinay; Powell, Jonathan J.; Kirkby, Karen J.; Grime, Geoffrey W.

2006-08-01

Microscopic particles (0.5-2 μm diameter), rich in calcium and phosphorus, are found in the lumen of the mid-distal gut of all mammals investigated, including humans, and these may play a role in immuno-surveillance and immune regulation of antigens from food and symbiotic bacteria that are contained in the gut. Whether these particles can cross in to tissue of the intestinal mucosa is unclear. If so, characterising their morphology and chemical composition is an important task in elucidating their function. The analysis of calcium phosphate in biological tissues has been approached in several ways including optical microscopy, scanning electron microscopy and, most recently in this work, with nuclear microscopy. In this paper, we describe the use of microPIXE and microRBS to locate these particles and to determine, accurately, the ratio of phosphorus to calcium using the information on sample thickness obtained from RBS to allow the PIXE ratios to be corrected. A commercial sample of hydroxy apatite was used to demonstrate accuracy and precision of the technique. Then, in a pilot study on intestinal tissue of mice, we demonstrated the presence of calcium phosphate microparticles, consistent with confocal microscopy observations, and we identified the average molar P:Ca molar ratio as 1.0. Further work will confirm the exact chemical speciation of these particles and will examine the influence of differing calcium containing diets on the formation of these microparticles.

Extended Field Laser Confocal Microscopy (EFLCM): Combining automated Gigapixel image capture with in silico virtual microscopy

PubMed Central

Flaberg, Emilie; Sabelström, Per; Strandh, Christer; Szekely, Laszlo

2008-01-01

Background Confocal laser scanning microscopy has revolutionized cell biology. However, the technique has major limitations in speed and sensitivity due to the fact that a single laser beam scans the sample, allowing only a few microseconds signal collection for each pixel. This limitation has been overcome by the introduction of parallel beam illumination techniques in combination with cold CCD camera based image capture. Methods Using the combination of microlens enhanced Nipkow spinning disc confocal illumination together with fully automated image capture and large scale in silico image processing we have developed a system allowing the acquisition, presentation and analysis of maximum resolution confocal panorama images of several Gigapixel size. We call the method Extended Field Laser Confocal Microscopy (EFLCM). Results We show using the EFLCM technique that it is possible to create a continuous confocal multi-colour mosaic from thousands of individually captured images. EFLCM can digitize and analyze histological slides, sections of entire rodent organ and full size embryos. It can also record hundreds of thousands cultured cells at multiple wavelength in single event or time-lapse fashion on fixed slides, in live cell imaging chambers or microtiter plates. Conclusion The observer independent image capture of EFLCM allows quantitative measurements of fluorescence intensities and morphological parameters on a large number of cells. EFLCM therefore bridges the gap between the mainly illustrative fluorescence microscopy and purely quantitative flow cytometry. EFLCM can also be used as high content analysis (HCA) instrument for automated screening processes. PMID:18627634

Quantitative tracking of tumor cells in phase-contrast microscopy exploiting halo artifact pattern

NASA Astrophysics Data System (ADS)

Kang, Mi-Sun; Song, Soo-Min; Lee, Hana; Kim, Myoung-Hee

2012-03-01

Tumor cell morphology is closely related to its invasiveness characteristics and migratory behaviors. An invasive tumor cell has a highly irregular shape, whereas a spherical cell is non-metastatic. Thus, quantitative analysis of cell features is crucial to determine tumor malignancy or to test the efficacy of anticancer treatment. We use phase-contrast microscopy to analyze single cell morphology and to monitor its change because it enables observation of long-term activity of living cells without photobleaching and phototoxicity, which is common in other fluorescence-labeled microscopy. Despite this advantage, there are image-level drawbacks to phase-contrast microscopy, such as local light effect and contrast interference ring, among others. Thus, we first applied a local filter to compensate for non-uniform illumination. Then, we used intensity distribution information to detect the cell boundary. In phase-contrast microscopy images, the cell normally appears as a dark region surrounded by a bright halo. As the halo artifact around the cell body is minimal and has an asymmetric diffusion pattern, we calculated the cross-sectional plane that intersected the center of each cell and was orthogonal to the first principal axis. Then, we extracted the dark cell region by level set. However, a dense population of cultured cells still rendered single-cell analysis difficult. Finally, we measured roundness and size to classify tumor cells into malignant and benign groups. We validated segmentation accuracy by comparing our findings with manually obtained results.

Extended Field Laser Confocal Microscopy (EFLCM): combining automated Gigapixel image capture with in silico virtual microscopy.

PubMed

Flaberg, Emilie; Sabelström, Per; Strandh, Christer; Szekely, Laszlo

2008-07-16

Confocal laser scanning microscopy has revolutionized cell biology. However, the technique has major limitations in speed and sensitivity due to the fact that a single laser beam scans the sample, allowing only a few microseconds signal collection for each pixel. This limitation has been overcome by the introduction of parallel beam illumination techniques in combination with cold CCD camera based image capture. Using the combination of microlens enhanced Nipkow spinning disc confocal illumination together with fully automated image capture and large scale in silico image processing we have developed a system allowing the acquisition, presentation and analysis of maximum resolution confocal panorama images of several Gigapixel size. We call the method Extended Field Laser Confocal Microscopy (EFLCM). We show using the EFLCM technique that it is possible to create a continuous confocal multi-colour mosaic from thousands of individually captured images. EFLCM can digitize and analyze histological slides, sections of entire rodent organ and full size embryos. It can also record hundreds of thousands cultured cells at multiple wavelength in single event or time-lapse fashion on fixed slides, in live cell imaging chambers or microtiter plates. The observer independent image capture of EFLCM allows quantitative measurements of fluorescence intensities and morphological parameters on a large number of cells. EFLCM therefore bridges the gap between the mainly illustrative fluorescence microscopy and purely quantitative flow cytometry. EFLCM can also be used as high content analysis (HCA) instrument for automated screening processes.

The three dimensionality of cell membranes: lamellar to cubic membrane transition as investigated by electron microscopy.

PubMed

Chong, Ketpin; Deng, Yuru

2012-01-01

Biological membranes are generally perceived as phospholipid bilayer structures that delineate in a lamellar form the cell surface and intracellular organelles. However, much more complex and highly convoluted membrane organizations are ubiquitously present in many cell types under certain types of stress, states of disease, or in the course of viral infections. Their occurrence under pathological conditions make such three-dimensionally (3D) folded and highly ordered membranes attractive biomarkers. They have also stimulated great biomedical interest in understanding the molecular basis of their formation. Currently, the analysis of such membrane arrangements, which include tubulo-reticular structures (TRS) or cubic membranes of various subtypes, is restricted to electron microscopic methods, including tomography. Preservation of membrane structures during sample preparation is the key to understand their true 3D nature. This chapter discusses methods for appropriate sample preparations to successfully examine and analyze well-preserved highly ordered membranes by electron microscopy. Processing methods and analysis conditions for green algae (Zygnema sp.) and amoeba (Chaos carolinense), mammalian cells in culture and primary tissue cells are described. We also discuss methods to identify cubic membranes by transmission electron microscopy (TEM) with the aid of a direct template matching method and by computer simulation. A 3D analysis of cubic cell membrane topology by electron tomography is described as well as scanning electron microscopy (SEM) to investigate surface contours of isolated mitochondria with cubic membrane arrangement. Copyright © 2012 Elsevier Inc. All rights reserved.

High-Precision Pinpointing of Luminescent Targets in Encoder-Assisted Scanning Microscopy Allowing High-Speed Quantitative Analysis.

PubMed

Zheng, Xianlin; Lu, Yiqing; Zhao, Jiangbo; Zhang, Yuhai; Ren, Wei; Liu, Deming; Lu, Jie; Piper, James A; Leif, Robert C; Liu, Xiaogang; Jin, Dayong

2016-01-19

Compared with routine microscopy imaging of a few analytes at a time, rapid scanning through the whole sample area of a microscope slide to locate every single target object offers many advantages in terms of simplicity, speed, throughput, and potential for robust quantitative analysis. Existing techniques that accommodate solid-phase samples incorporating individual micrometer-sized targets generally rely on digital microscopy and image analysis, with intrinsically low throughput and reliability. Here, we report an advanced on-the-fly stage scanning method to achieve high-precision target location across the whole slide. By integrating X- and Y-axis linear encoders to a motorized stage as the virtual "grids" that provide real-time positional references, we demonstrate an orthogonal scanning automated microscopy (OSAM) technique which can search a coverslip area of 50 × 24 mm(2) in just 5.3 min and locate individual 15 μm lanthanide luminescent microspheres with standard deviations of 1.38 and 1.75 μm in X and Y directions. Alongside implementation of an autofocus unit that compensates the tilt of a slide in the Z-axis in real time, we increase the luminescence detection efficiency by 35% with an improved coefficient of variation. We demonstrate the capability of advanced OSAM for robust quantification of luminescence intensities and lifetimes for a variety of micrometer-scale luminescent targets, specifically single down-shifting and upconversion microspheres, crystalline microplates, and color-barcoded microrods, as well as quantitative suspension array assays of biotinylated-DNA functionalized upconversion nanoparticles.

Chemical Mapping of Essential Oils, Flavonoids and Carotenoids in Citrus Peels by Raman Microscopy.

PubMed

Yang, Ying; Wang, Xiaohe; Zhao, Chengying; Tian, Guifang; Zhang, Hua; Xiao, Hang; He, Lili; Zheng, Jinkai

2017-12-01

Citrus peels, by-products in large quantity, are rich in various functional and beneficial components which have wide applications. Chemical analysis of these components in citrus peels is an important step to determine the usefulness of the by-products for further applications. In this study, we explored Raman microscopy for rapid, nondestructive, and in situ chemical mapping of multiple main functional components from citrus peels. The relative amount and distribution in different locations (flavedo, albedo, and longitudinal section) of 3 main functional components (essential oils, carotenoids, and flavonoids) in citrus peels were systematically investigated. The distribution profiles of these components were heterogeneous on the peels and varied between different species of citrus peels. Essential oil was found mainly existed in the oil glands, while carotenoids were in the complementary location. Some flavonoids were observed in the oil glands. This study showed the capability of Raman microscopy for rapid and nondestructive analysis of multiple bio-components without extraction from plants. The information obtained from this study would assist the better production and application of the functional and beneficial components from citrus by products in an effective and sustainable manner. This study indicated the capability of Raman microscopy for rapid and nondestructive analysis of multiple bioactive components in plant tissues. The information obtained from the study would be valuable for developing effective and sustainable strategy of utilization of citrus peels for further applications. © 2017 Institute of Food Technologists®.

The Electron Microscopy Outreach Program: A Web-based resource for research and education.

PubMed

Sosinsky, G E; Baker, T S; Hand, G; Ellisman, M H

1999-01-01

We have developed a centralized World Wide Web (WWW)-based environment that serves as a resource of software tools and expertise for biological electron microscopy. A major focus is molecular electron microscopy, but the site also includes information and links on structural biology at all levels of resolution. This site serves to help integrate or link structural biology techniques in accordance with user needs. The WWW site, called the Electron Microscopy (EM) Outreach Program (URL: http://emoutreach.sdsc.edu), provides scientists with computational and educational tools for their research and edification. In particular, we have set up a centralized resource containing course notes, references, and links to image analysis and three-dimensional reconstruction software for investigators wanting to learn about EM techniques either within or outside of their fields of expertise. Copyright 1999 Academic Press.

Atomic force microscopy imaging of fragments from the Martian meteorite ALH84001

NASA Technical Reports Server (NTRS)

Steele, A.; Goddard, D.; Beech, I. B.; Tapper, R. C.; Stapleton, D.; Smith, J. R.

1998-01-01

A combination of scanning electron microscopy (SEM) and environmental scanning electron microscopy (ESEM) techniques, as well as atomic force microscopy (AFM) methods has been used to study fragments of the Martian meteorite ALH84001. Images of the same areas on the meteorite were obtained prior to and following gold/palladium coating by mapping the surface of the fragment using ESEM coupled with energy-dispersive X-ray analysis. Viewing of the fragments demonstrated the presence of structures, previously described as nanofossils by McKay et al. (Search for past life on Mars--possible relic biogenic activity in martian meteorite ALH84001. Science, 1996, pp. 924-930) of NASA who used SEM imaging of gold-coated meteorite samples. Careful imaging of the fragments revealed that the observed structures were not an artefact introduced by the coating procedure.

On the state of crystallography at the dawn of the electron microscopy revolution.

PubMed

Higgins, Matthew K; Lea, Susan M

2017-10-01

While protein crystallography has, for many years, been the most used method for structural analysis of macromolecular complexes, remarkable recent advances in high-resolution electron cryo-microscopy led to suggestions that 'the revolution will not be crystallised'. Here we highlight the current success rate, speed and ease of modern crystallographic structure determination and some recent triumphs of both 'classical' crystallography and the use of X-ray free electron lasers. We also outline fundamental differences between structure determination using X-ray crystallography and electron microscopy. We suggest that crystallography will continue to co-exist with electron microscopy as part of an integrated array of methods, allowing structural biologists to focus on fundamental biological questions rather than being constrained by the methods available. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Non-rigid multi-frame registration of cell nuclei in live cell fluorescence microscopy image data.

PubMed

Tektonidis, Marco; Kim, Il-Han; Chen, Yi-Chun M; Eils, Roland; Spector, David L; Rohr, Karl

2015-01-01

The analysis of the motion of subcellular particles in live cell microscopy images is essential for understanding biological processes within cells. For accurate quantification of the particle motion, compensation of the motion and deformation of the cell nucleus is required. We introduce a non-rigid multi-frame registration approach for live cell fluorescence microscopy image data. Compared to existing approaches using pairwise registration, our approach exploits information from multiple consecutive images simultaneously to improve the registration accuracy. We present three intensity-based variants of the multi-frame registration approach and we investigate two different temporal weighting schemes. The approach has been successfully applied to synthetic and live cell microscopy image sequences, and an experimental comparison with non-rigid pairwise registration has been carried out. Copyright © 2014 Elsevier B.V. All rights reserved.

New approaches for the analysis of confluent cell layers with quantitative phase digital holographic microscopy

NASA Astrophysics Data System (ADS)

Pohl, L.; Kaiser, M.; Ketelhut, S.; Pereira, S.; Goycoolea, F.; Kemper, Björn

2016-03-01

Digital holographic microscopy (DHM) enables high resolution non-destructive inspection of technical surfaces and minimally-invasive label-free live cell imaging. However, the analysis of confluent cell layers represents a challenge as quantitative DHM phase images in this case do not provide sufficient information for image segmentation, determination of the cellular dry mass or calculation of the cell thickness. We present novel strategies for the analysis of confluent cell layers with quantitative DHM phase contrast utilizing a histogram based-evaluation procedure. The applicability of our approach is illustrated by quantification of drug induced cell morphology changes and it is shown that the method is capable to quantify reliable global morphology changes of confluent cell layers.

Quantitative analysis of phosphoinositide 3-kinase (PI3K) signaling using live-cell total internal reflection fluorescence (TIRF) microscopy.

PubMed

Johnson, Heath E; Haugh, Jason M

2013-12-02

This unit focuses on the use of total internal reflection fluorescence (TIRF) microscopy and image analysis methods to study the dynamics of signal transduction mediated by class I phosphoinositide 3-kinases (PI3Ks) in mammalian cells. The first four protocols cover live-cell imaging experiments, image acquisition parameters, and basic image processing and segmentation. These methods are generally applicable to live-cell TIRF experiments. The remaining protocols outline more advanced image analysis methods, which were developed in our laboratory for the purpose of characterizing the spatiotemporal dynamics of PI3K signaling. These methods may be extended to analyze other cellular processes monitored using fluorescent biosensors. Copyright © 2013 John Wiley & Sons, Inc.

Determination of element composition and extraterrestrial material occurrence in moss and lichen samples from King George Island (Antarctica) using reactor neutron activation analysis and SEM microscopy.

PubMed

Mróz, Tomasz; Szufa, Katarzyna; Frontasyeva, Marina V; Tselmovich, Vladimir; Ostrovnaya, Tatiana; Kornaś, Andrzej; Olech, Maria A; Mietelski, Jerzy W; Brudecki, Kamil

2018-01-01

Seven lichens (Usnea antarctica and U. aurantiacoatra) and nine moss samples (Sanionia uncinata) collected in King George Island were analyzed using instrumental neutron activation analysis, and concentration of major and trace elements was calculated. For some elements, the concentrations observed in moss samples were higher than corresponding values reported from other sites in the Antarctica, but in the lichens, these were in the same range of concentrations. Scanning electron microscopy (SEM) and statistical analysis showed large influence of volcanic-origin particles. Also, the interplanetary cosmic particles (ICP) were observed in investigated samples, as mosses and lichens are good collectors of ICP and micrometeorites.

Biocomposite Plasma-Sprayed Coatings Based on Zinc-Substituted Hydroxyapatite: Structure, Properties, and Prospects of Application

NASA Astrophysics Data System (ADS)

Lyasnikova, A. V.; Markelova, O. A.; Lyasnikov, V. N.; Dudareva, O. A.

2016-01-01

The method of synthesis of a zinc-substituted hydroxyapatite powder is presented, and the technology of creating coatings by its spraying is described. The results of studies on the morphological, physical, and chemical parameters of a zinc-substituted hydroxyapatite coating by using X-ray analysis, infrared spectroscopy, transmission electron microscopy, optical microscopy, SEM, and other methods are given.

Analysis of electromagnetic forces and causality in electron microscopy.

PubMed

Reyes-Coronado, Alejandro; Ortíz-Solano, Carlos Gael; Zabala, Nerea; Rivacoba, Alberto; Esquivel-Sirvent, Raúl

2018-09-01

The non-physical effects on the transverse momentum transfer from fast electrons to gold nanoparticles associated to the use of non-causal dielectric functions are studied. A direct test of the causality based on the surface Kramers-Kronig relations is presented. This test is applied to the different dielectric function used to describe gold nanostructures in electron microscopy. Copyright © 2018. Published by Elsevier B.V.

Stochastic optical reconstruction microscopy-based relative localization analysis (STORM-RLA) for quantitative nanoscale assessment of spatial protein organization.

PubMed

Veeraraghavan, Rengasayee; Gourdie, Robert G

2016-11-07

The spatial association between proteins is crucial to understanding how they function in biological systems. Colocalization analysis of fluorescence microscopy images is widely used to assess this. However, colocalization analysis performed on two-dimensional images with diffraction-limited resolution merely indicates that the proteins are within 200-300 nm of each other in the xy-plane and within 500-700 nm of each other along the z-axis. Here we demonstrate a novel three-dimensional quantitative analysis applicable to single-molecule positional data: stochastic optical reconstruction microscopy-based relative localization analysis (STORM-RLA). This method offers significant advantages: 1) STORM imaging affords 20-nm resolution in the xy-plane and <50 nm along the z-axis; 2) STORM-RLA provides a quantitative assessment of the frequency and degree of overlap between clusters of colabeled proteins; and 3) STORM-RLA also calculates the precise distances between both overlapping and nonoverlapping clusters in three dimensions. Thus STORM-RLA represents a significant advance in the high-throughput quantitative assessment of the spatial organization of proteins. © 2016 Veeraraghavan and Gourdie. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Automated fluorescent miscroscopic image analysis of PTBP1 expression in glioma

PubMed Central

Becker, Aline; Elder, Brad; Puduvalli, Vinay; Winter, Jessica; Gurcan, Metin

2017-01-01

Multiplexed immunofluorescent testing has not entered into diagnostic neuropathology due to the presence of several technical barriers, amongst which includes autofluorescence. This study presents the implementation of a methodology capable of overcoming the visual challenges of fluorescent microscopy for diagnostic neuropathology by using automated digital image analysis, with long term goal of providing unbiased quantitative analyses of multiplexed biomarkers for solid tissue neuropathology. In this study, we validated PTBP1, a putative biomarker for glioma, and tested the extent to which immunofluorescent microscopy combined with automated and unbiased image analysis would permit the utility of PTBP1 as a biomarker to distinguish diagnostically challenging surgical biopsies. As a paradigm, we utilized second resections from patients diagnosed either with reactive brain changes (pseudoprogression) and recurrent glioblastoma (true progression). Our image analysis workflow was capable of removing background autofluorescence and permitted quantification of DAPI-PTBP1 positive cells. PTBP1-positive nuclei, and the mean intensity value of PTBP1 signal in cells. Traditional pathological interpretation was unable to distinguish between groups due to unacceptably high discordance rates amongst expert neuropathologists. Our data demonstrated that recurrent glioblastoma showed more DAPI-PTBP1 positive cells and a higher mean intensity value of PTBP1 signal compared to resections from second surgeries that showed only reactive gliosis. Our work demonstrates the potential of utilizing automated image analysis to overcome the challenges of implementing fluorescent microscopy in diagnostic neuropathology. PMID:28282372

Implementation of a National Reference Laboratory for Buruli Ulcer Disease in Togo

PubMed Central

Badziklou, Kossi; Halatoko, Wemboo Afiwa; Maman, Issaka; Vogel, Felix; Bidjada, Bawimodom; Awoussi, Koffi Somenou; Piten, Ebekalisai; Helfrich, Kerstin; Mengele, Carolin; Nitschke, Jörg; Amekuse, Komi; Wiedemann, Franz Xaver; Diefenhardt, Adolf; Kobara, Basile; Herbinger, Karl–Heinz; Kere, Abiba Banla; Prince-David, Mireille; Löscher, Thomas; Bretzel, Gisela

2013-01-01

Background In a previous study PCR analysis of clinical samples from suspected cases of Buruli ulcer disease (BUD) from Togo and external quality assurance (EQA) for local microscopy were conducted at an external reference laboratory in Germany. The relatively poor performance of local microscopy as well as effort and time associated with shipment of PCR samples necessitated the implementation of stringent EQA measures and availability of local laboratory capacity. This study describes the approach to implementation of a national BUD reference laboratory in Togo. Methodology Large scale outreach activities accompanied by regular training programs for health care professionals were conducted in the regions “Maritime” and “Central,” standard operating procedures defined all processes in participating laboratories (regional, national and external reference laboratories) as well as the interaction between laboratories and partners in the field. Microscopy was conducted at regional level and slides were subjected to EQA at national and external reference laboratories. For PCR analysis, sample pairs were collected and subjected to a dry-reagent-based IS2404-PCR (DRB-PCR) at national level and standard IS2404 PCR followed by IS2404 qPCR analysis of negative samples at the external reference laboratory. Principal Findings The inter-laboratory concordance rates for microscopy ranged from 89% to 94%; overall, microscopy confirmed 50% of all suspected BUD cases. The inter-laboratory concordance rate for PCR was 96% with an overall PCR case confirmation rate of 78%. Compared to a previous study, the rate of BUD patients with non-ulcerative lesions increased from 37% to 50%, the mean duration of disease before clinical diagnosis decreased significantly from 182.6 to 82.1 days among patients with ulcerative lesions, and the percentage of category III lesions decreased from 30.3% to 19.2%. Conclusions High inter-laboratory concordance rates as well as case confirmation rates of 50% (microscopy), 71% (PCR at national level), and 78% (including qPCR confirmation at external reference laboratory) suggest high standards of BUD diagnostics. The increase of non-ulcerative lesions, as well as the decrease in diagnostic delay and category III lesions, prove the effect of comprehensive EQA and training measures involving also procedures outside the laboratory. PMID:23359828

Implementation of a national reference laboratory for Buruli ulcer disease in Togo.

PubMed

Beissner, Marcus; Huber, Kristina Lydia; Badziklou, Kossi; Halatoko, Wemboo Afiwa; Maman, Issaka; Vogel, Felix; Bidjada, Bawimodom; Awoussi, Koffi Somenou; Piten, Ebekalisai; Helfrich, Kerstin; Mengele, Carolin; Nitschke, Jörg; Amekuse, Komi; Wiedemann, Franz Xaver; Diefenhardt, Adolf; Kobara, Basile; Herbinger, Karl-Heinz; Kere, Abiba Banla; Prince-David, Mireille; Löscher, Thomas; Bretzel, Gisela

2013-01-01

In a previous study PCR analysis of clinical samples from suspected cases of Buruli ulcer disease (BUD) from Togo and external quality assurance (EQA) for local microscopy were conducted at an external reference laboratory in Germany. The relatively poor performance of local microscopy as well as effort and time associated with shipment of PCR samples necessitated the implementation of stringent EQA measures and availability of local laboratory capacity. This study describes the approach to implementation of a national BUD reference laboratory in Togo. Large scale outreach activities accompanied by regular training programs for health care professionals were conducted in the regions "Maritime" and "Central," standard operating procedures defined all processes in participating laboratories (regional, national and external reference laboratories) as well as the interaction between laboratories and partners in the field. Microscopy was conducted at regional level and slides were subjected to EQA at national and external reference laboratories. For PCR analysis, sample pairs were collected and subjected to a dry-reagent-based IS2404-PCR (DRB-PCR) at national level and standard IS2404 PCR followed by IS2404 qPCR analysis of negative samples at the external reference laboratory. The inter-laboratory concordance rates for microscopy ranged from 89% to 94%; overall, microscopy confirmed 50% of all suspected BUD cases. The inter-laboratory concordance rate for PCR was 96% with an overall PCR case confirmation rate of 78%. Compared to a previous study, the rate of BUD patients with non-ulcerative lesions increased from 37% to 50%, the mean duration of disease before clinical diagnosis decreased significantly from 182.6 to 82.1 days among patients with ulcerative lesions, and the percentage of category III lesions decreased from 30.3% to 19.2%. High inter-laboratory concordance rates as well as case confirmation rates of 50% (microscopy), 71% (PCR at national level), and 78% (including qPCR confirmation at external reference laboratory) suggest high standards of BUD diagnostics. The increase of non-ulcerative lesions, as well as the decrease in diagnostic delay and category III lesions, prove the effect of comprehensive EQA and training measures involving also procedures outside the laboratory.

Rapid detection of biofilms and adherent pathogens using scanning confocal laser microscopy and episcopic differential interference contrast microscopy.

PubMed

Keevil, C W

2003-01-01

Knowledge of biofilm structure and function has changed significantly in the last few years due to advances in light microscopy. One pertinent example is the use of scanning confocal laser microscopy (SCLM) to visualise corrosion pits caused by the biofilm mosaic footprint on corroding metal surfaces. Nevertheless, SCLM has some limitations as to its widespread use, including cost, inability to observe motile bacteria and eukaryotic grazers within biofilms, and difficulty to scan a curved surface. By contrast, episcopic differential interference contrast (EDIC) microscopy has provided a rapid, real time analysis of biofilms on opaque, curved, natural or man-made surfaces without the need for cover slips and oil. EDIC, coupled with epi-fluorescence (EDIC/EF), microscopy has been used successfully to visualise the 3-D biofilm structure, physiological niches, protozoal grazing and iron biomineralization, and the location of specific pathogens such as Legionella pneumophila, Campylobacter jejuni and Cryptosporidium parvum. These species were identified using gold nanoparticles or fluorophores coupled to monoclonal antibodies or 16S rRNA probes, respectively. Among its many potential uses, the EDIC technique will provide a rapid procedure to facilitate the calibration of the modern generation of biofilm-sensing electrodes.

Accessible microscopy workstation for students and scientists with mobility impairments.

PubMed

Duerstock, Bradley S

2006-01-01

An integrated accessible microscopy workstation was designed and developed to allow persons with mobility impairments to control all aspects of light microscopy with minimal human assistance. This system, named AccessScope, is capable of performing brightfield and fluorescence microscopy, image analysis, and tissue morphometry requisite for undergraduate science courses to graduate-level research. An accessible microscope is necessary for students and scientists with mobility impairments to be able to use a microscope independently to better understand microscopical imaging concepts and cell biology. This knowledge is not always apparent by simply viewing a catalog of histological images. The ability to operate a microscope independently eliminates the need to hire an assistant or rely on a classmate and permits one to take practical laboratory examinations by oneself. Independent microscope handling is also crucial for graduate students and scientists with disabilities to perform scientific research. By making a personal computer as the user interface for controlling AccessScope functions, different upper limb mobility impairments could be accommodated by using various computer input devices and assistive technology software. Participants with a range of upper limb mobility impairments evaluated the prototype microscopy workstation. They were able to control all microscopy functions including loading different slides without assistance.

Virtual microscopy and digital pathology in training and education.

PubMed

Hamilton, Peter W; Wang, Yinhai; McCullough, Stephen J

2012-04-01

Traditionally, education and training in pathology has been delivered using textbooks, glass slides and conventional microscopy. Over the last two decades, the number of web-based pathology resources has expanded dramatically with centralized pathological resources being delivered to many students simultaneously. Recently, whole slide imaging technology allows glass slides to be scanned and viewed on a computer screen via dedicated software. This technology is referred to as virtual microscopy and has created enormous opportunities in pathological training and education. Students are able to learn key histopathological skills, e.g. to identify areas of diagnostic relevance from an entire slide, via a web-based computer environment. Students no longer need to be in the same room as the slides. New human-computer interfaces are also being developed using more natural touch technology to enhance the manipulation of digitized slides. Several major initiatives are also underway introducing online competency and diagnostic decision analysis using virtual microscopy and have important future roles in accreditation and recertification. Finally, researchers are investigating how pathological decision-making is achieved using virtual microscopy and modern eye-tracking devices. Virtual microscopy and digital pathology will continue to improve how pathology training and education is delivered. © 2012 The Authors APMIS © 2012 APMIS.

Induction of morphological changes in death-induced cancer cells monitored by holographic microscopy.

PubMed

El-Schich, Zahra; Mölder, Anna; Tassidis, Helena; Härkönen, Pirkko; Falck Miniotis, Maria; Gjörloff Wingren, Anette

2015-03-01

We are using the label-free technique of holographic microscopy to analyze cellular parameters including cell number, confluence, cellular volume and area directly in the cell culture environment. We show that death-induced cells can be distinguished from untreated counterparts by the use of holographic microscopy, and we demonstrate its capability for cell death assessment. Morphological analysis of two representative cell lines (L929 and DU145) was performed in the culture flasks without any prior cell detachment. The two cell lines were treated with the anti-tumour agent etoposide for 1-3days. Measurements by holographic microscopy showed significant differences in average cell number, confluence, volume and area when comparing etoposide-treated with untreated cells. The cell volume of the treated cell lines was initially increased at early time-points. By time, cells decreased in volume, especially when treated with high doses of etoposide. In conclusion, we have shown that holographic microscopy allows label-free and completely non-invasive morphological measurements of cell growth, viability and death. Future applications could include real-time monitoring of these holographic microscopy parameters in cells in response to clinically relevant compounds. Copyright © 2015 Elsevier Inc. All rights reserved.

Biomimetic novel nanoporous niobium oxide coating for orthopaedic applications

NASA Astrophysics Data System (ADS)

Pauline, S. Anne; Rajendran, N.

2014-01-01

Niobium oxide was synthesized by sol-gel methodology and a crystalline, nanoporous and adherent coating of Nb2O5 was deposited on 316L SS using the spin coating technique and heat treatment. The synthesis conditions were optimized to obtain a nanoporous morphology. The coating was characterized using attenuated total reflectance-Infrared spectroscopy (ATR-IR), X-ray diffraction analysis (XRD), scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDX), atomic force microscopy (AFM) and transmission electron microscopy (TEM) and the formation of crystalline Nb2O5 coating with nanoporous morphology was confirmed. Mechanical studies confirmed that the coating has excellent adherence to the substrate and the hardness value of the coating was excellent. Contact angle analysis showed increased hydrophilicity for the coated substrate. In vitro bioactivity test confirmed that the Nb2O5 coating with nanoporous morphology facilitated the growth of hydroxyapatite (HAp). This was further confirmed by the solution analysis test where increased uptake of calcium and phosphorous ions from simulated body fluid (SBF) was observed. Electrochemical evaluation of the coating confirmed that the crystalline coating is insulative and protective in nature and offered excellent corrosion protection to 316L SS. Thus, this study confirmed that the nanoporous crystalline Nb2O5 coating conferred bioactivity and enhanced corrosion resistance on 316L SS.

Automated analysis of individual sperm cells using stain-free interferometric phase microscopy and machine learning.

PubMed

Mirsky, Simcha K; Barnea, Itay; Levi, Mattan; Greenspan, Hayit; Shaked, Natan T

2017-09-01

Currently, the delicate process of selecting sperm cells to be used for in vitro fertilization (IVF) is still based on the subjective, qualitative analysis of experienced clinicians using non-quantitative optical microscopy techniques. In this work, a method was developed for the automated analysis of sperm cells based on the quantitative phase maps acquired through use of interferometric phase microscopy (IPM). Over 1,400 human sperm cells from 8 donors were imaged using IPM, and an algorithm was designed to digitally isolate sperm cell heads from the quantitative phase maps while taking into consideration both the cell 3D morphology and contents, as well as acquire features describing sperm head morphology. A subset of these features was used to train a support vector machine (SVM) classifier to automatically classify sperm of good and bad morphology. The SVM achieves an area under the receiver operating characteristic curve of 88.59% and an area under the precision-recall curve of 88.67%, as well as precisions of 90% or higher. We believe that our automatic analysis can become the basis for objective and automatic sperm cell selection in IVF. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

Blood pulse wave velocity measured by photoacoustic microscopy

NASA Astrophysics Data System (ADS)

Yeh, Chenghung; Hu, Song; Maslov, Konstantin; Wang, Lihong V.

2013-03-01

Blood pulse wave velocity (PWV) is an important indicator for vascular stiffness. In this letter, we present electrocardiogram-synchronized photoacoustic microscopy for in vivo noninvasive quantification of the PWV in the peripheral vessels of mice. Interestingly, strong correlation between blood flow speed and ECG were clearly observed in arteries but not in veins. PWV is measured by the pulse travel time and the distance between two spot of a chose vessel, where simultaneously recorded electrocardiograms served as references. Statistical analysis shows a linear correlation between the PWV and the vessel diameter, which agrees with known physiology. Keywords: photoacoustic microscopy, photoacoustic spectroscopy, bilirubin, scattering medium.

Room temperature chemical synthesis of lead selenide thin films with preferred orientation

NASA Astrophysics Data System (ADS)

Kale, R. B.; Sartale, S. D.; Ganesan, V.; Lokhande, C. D.; Lin, Yi-Feng; Lu, Shih-Yuan

2006-11-01

Room temperature chemical synthesis of PbSe thin films was carried out from aqueous ammoniacal solution using Pb(CH3COO)2 as Pb2+ and Na2SeSO3 as Se2- ion sources. The films were characterized by a various techniques including, X-ray diffraction (XRD), energy dispersive X-ray analysis (EDAX), scanning electron microscopy (SEM), transmission electron microscopy (TEM), high resolution transmission electron microscopy (HR-TEM), selected area electron diffraction (SAED), Fast Fourier transform (FFT) and UV-vis-NIR techniques. The study revealed that the PbSe thin film consists of preferentially oriented nanocubes with energy band gap of 0.5 eV.

Nanocomposite of polyaniline nanorods grown on graphene nanoribbons for highly capacitive pseudocapacitors.

PubMed

Li, Lei; Raji, Abdul-Rahman O; Fei, Huilong; Yang, Yang; Samuel, Errol L G; Tour, James M

2013-07-24

A facile and cost-effective approach to the fabrication of a nanocomposite material of polyaniline (PANI) and graphene nanoribbons (GNRs) has been developed. The morphology of the composite was characterized by scanning electron microscopy, transmission electron microscopy, X-ray photoelectron microscopy, and X-ray diffraction analysis. The resulting composite has a high specific capacitance of 340 F/g and stable cycling performance with 90% capacitance retention over 4200 cycles. The high performance of the composite results from the synergistic combination of electrically conductive GNRs and highly capacitive PANI. The method developed here is practical for large-scale development of pseudocapacitor electrodes for energy storage.

Analytical electron microscopy of Mg-SiO smokes - A comparison with infrared and XRD studies

NASA Technical Reports Server (NTRS)

Rietmeijer, F. J. M.; Nuth, J. A.; Mackinnon, I. D. R.

1986-01-01

Analytical electron microscopy conducted for Mg-SiO smokes (experimentally obtained from samples previously characterized by IR spectroscopy) indicates that the microcrystallinity content of unannealed smokes increases with increased annealing for up to 30 hr. The growth of forsterite microcrystallites in the initially nonstoichiometric smokes may give rise to the contemporaneous growth of the SiO polymorph tridymite and MgO; after 4 hr of annealing, these react to form enstatite. It is suggested that XRD analysis and IR spectroscopy should be conducted in conjunction with detailed analytical electron microscopy for the detection of emerging crystallinity in vapor-phase condensates.

Confocal microscopy to guide laser ablation of basal cell carinoma: a preliminary feasibility study

NASA Astrophysics Data System (ADS)

Larson, Bjorg A.; Sierra, Heidy; Chen, Jason; Rajadhyaksha, Milind

2013-03-01

Laser ablation may be a promising method for removal of skin lesions, with the potential for better cosmetic outcomes and reduced scarring and infection. An obstacle to implementing laser ablation is that the treatment leaves no tissue for histopathological analysis. Pre-operative and intra-operative mapping of BCCs using confocal microscopy may guide the ablation of the tumor until all tumor is removed. We demonstrate preliminary feasibility of confocal microscopy to guide laser ablation of BCCs in freshly excised tissue from Mohs surgery. A 2940 nm Er:YAG laser provides efficient ablation of tumor with reduced thermal damage to the surrounding tissue.

Measuring skin penetration by confocal Raman microscopy (CRM): correlation to results from conventional experiments

NASA Astrophysics Data System (ADS)

Lunter, Dominique; Daniels, Rolf

2016-03-01

Confocal Raman microscopy has become an advancing technique in the characterization of drug transport into the skin. In this study the skin penetration of a local anesthetic from a semisolid preparation was investigated. Furthermore, the effect of the chemical enhancers propylene glycol and POE-23-lauryl ether on its penetration was investigated. The results show that confocal Raman microscopy may provide detailed information on the penetration of APIs into the skin and may elucidate their distribution within the skin with high resolution. The results of the CRM analysis are fully in line with those of conventional permeation and penetration experiments.

25 years of telepathology research: a bibliometric analysis.

PubMed

Della Mea, Vincenzo

2011-03-30

The first appearance of the word "telepathology" in a scientific paper can be tracked down to 1986, in a famous editorial of Ronald Weinstein. Since that paper, research in telepathology grew up developing different subfields, including static and dynamic telepathology and more recently virtual microscopy. The present work attempts an analysis of research in telepathology, starting from the tools provided by bibliometrics. A query has been developed to extract papers related to telepathology and virtual microscopy, and it has been then submitted to Pubmed by means of Entrez Utilities functions. Results obtained in XML have been processed through ad-hoc developed PHP scripts, in order to extract data on Authors, countries, and keywords. On PubMed, 967 papers related to telepathology and virtual microscopy have been retrieved, which involved 2904 Authors; corresponding authors were from 37 countries. Of those authors, 2213 co-authored just one paper. Papers were published on 344 different journals, of which only 52 from the Pathology field. An analysis of papers per year has been also attempted, that demonstrates variable research output in time. From the proposed analysis, telepathology seems to have been consistently studied, in time, by about 400 researchers, with occasional participation of many other people. Telepathology research seems also to have varied in time, although some peaks in paper publishing are certainly related to the proceedings of the European congress on telepathology series, when they have been published on journals. However, some clear sign appears that suggests research in traditional telepathology, after a peak in 2000, showed some decline until virtual microscopy became mainstream, topic that currently pushes research again. The low number of clinical trials calls for more randomized studies in telepathology, to enable evidence-based application.

25 years of telepathology research: a bibliometric analysis

PubMed Central

2011-01-01

Background The first appearance of the word “telepathology” in a scientific paper can be tracked down to 1986, in a famous editorial of Ronald Weinstein. Since that paper, research in telepathology grew up developing different subfields, including static and dynamic telepathology and more recently virtual microscopy. The present work attempts an analysis of research in telepathology, starting from the tools provided by bibliometrics. Methods A query has been developed to extract papers related to telepathology and virtual microscopy, and it has been then submitted to Pubmed by means of Entrez Utilities functions. Results obtained in XML have been processed through ad-hoc developed PHP scripts, in order to extract data on Authors, countries, and keywords. Results On PubMed, 967 papers related to telepathology and virtual microscopy have been retrieved, which involved 2904 Authors; corresponding authors were from 37 countries. Of those authors, 2213 co-authored just one paper. Papers were published on 344 different journals, of which only 52 from the Pathology field. An analysis of papers per year has been also attempted, that demonstrates variable research output in time. Conclusions From the proposed analysis, telepathology seems to have been consistently studied, in time, by about 400 researchers, with occasional participation of many other people. Telepathology research seems also to have varied in time, although some peaks in paper publishing are certainly related to the proceedings of the European congress on telepathology series, when they have been published on journals. However, some clear sign appears that suggests research in traditional telepathology, after a peak in 2000, showed some decline until virtual microscopy became mainstream, topic that currently pushes research again. The low number of clinical trials calls for more randomized studies in telepathology, to enable evidence-based application. PMID:21489197

Detection of Onchocerca volvulus in Skin Snips by Microscopy and Real-Time Polymerase Chain Reaction: Implications for Monitoring and Evaluation Activities.

PubMed

Thiele, Elizabeth A; Cama, Vitaliano A; Lakwo, Thomson; Mekasha, Sindeaw; Abanyie, Francisca; Sleshi, Markos; Kebede, Amha; Cantey, Paul T

2016-04-01

Microscopic evaluation of skin biopsies is the monitoring and evaluation (M and E) method currently used by multiple onchocerciasis elimination programs in Africa. However, as repeated mass drug administration suppresses microfilarial loads, the sensitivity and programmatic utility of skin snip microscopy is expected to decrease. Using a pan-filarial real-time polymerase chain reaction with melt curve analysis (qPCR-MCA), we evaluated 1) the use of a single-step molecular assay for detecting and identifying Onchocerca volvulus microfilariae in residual skin snips and 2) the sensitivity of skin snip microscopy relative to qPCR-MCA. Skin snips were collected and examined with routine microscopy in hyperendemic regions of Uganda and Ethiopia (N= 500 each) and "residual" skin snips (tissue remaining after induced microfilarial emergence) were tested with qPCR-MCA. qPCR-MCA detected Onchocerca DNA in 223 residual snips: 139 of 147 microscopy(+) and 84 among microscopy(-) snips, suggesting overall sensitivity of microscopy was 62.3% (139/223) relative to qPCR-MCA (75.6% in Uganda and 28.6% in Ethiopia). These findings demonstrate the insufficient sensitivity of skin snip microscopy for reliable programmatic monitoring. Molecular tools such as qPCR-MCA can augment sensitivity and provide diagnostic confirmation of skin biopsies and will be useful for evaluation or validation of new onchocerciasis M and E tools. © The American Society of Tropical Medicine and Hygiene.

A review of advantages of high-efficiency X-ray spectrum imaging for analysis of nanostructured ferritic alloys

DOE PAGES

Parish, Chad M.; Miller, Michael K.

2014-12-09

Nanostructured ferritic alloys (NFAs) exhibit complex microstructures consisting of 100-500 nm ferrite grains, grain boundary solute enrichment, and multiple populations of precipitates and nanoclusters (NCs). Understanding these materials' excellent creep and radiation-tolerance properties requires a combination of multiple atomic-scale experimental techniques. Recent advances in scanning transmission electron microscopy (STEM) hardware and data analysis methods have the potential to revolutionize nanometer to micrometer scale materials analysis. The application of these methods is applied to NFAs as a test case and is compared to both conventional STEM methods as well as complementary methods such as scanning electron microscopy and atom probe tomography.more » In this paper, we review past results and present new results illustrating the effectiveness of latest-generation STEM instrumentation and data analysis.« less

Electron Microscopy Abrasion Analysis of Candidate Fabrics for Planetary Space Suit Protective Overgarment Application

NASA Technical Reports Server (NTRS)

Hennessy, Mary J.

1992-01-01

The Electron Microscopy Abrasion Analysis of Candidate Fabrics for Planetary Space Suit Protective Overgarment Application is in support of the Abrasion Resistance Materials Screening Test. The fundamental assumption made for the SEM abrasion analysis was that woven fabrics to be used as the outermost layer of the protective overgarment in the design of the future, planetary space suits perform best when new. It is the goal of this study to determine which of the candidate fabrics was abraded the least in the tumble test. The sample that was abraded the least will be identified at the end of the report as the primary candidate fabric for further investigation. In addition, this analysis will determine if the abrasion seen by the laboratory tumbled samples is representative of actual EVA Apollo abrasion.

An Integrated Approach to Thermal Analysis of Pharmaceutical Solids

ERIC Educational Resources Information Center

Riley, Shelley R. Rabel

2015-01-01

A three-tiered experiment for undergraduate Instrumental Analysis students is presented in which students characterize the solid-state thermal behavior of an active pharmaceutical ingredient (acetaminophen) and excipient (a-lactose hydrate) using differential scanning calorimetry, thermogravimetric analysis, and thermal microscopy. Students are…

Introduction: A Symposium in Honor of Professor Sir John Meurig Thomas

NASA Astrophysics Data System (ADS)

Gai, P. L.; Saka, H.; Tomokiyo, Y.; Boyes, E. D.

2002-02-01

This issue is dedicated to Professor Sir John Meurig Thomas for his renowned contributions to electron microscopy in the chemical sciences. It is a collection of peer-reviewed leading articles in electron microscopy, based on the presentations at the Microscopy and Microanalysis (M&M) 2000 symposium, which was held to honor Professor Thomas's exceptional scientific leadership and wide-ranging fundamental contributions in the chemical applications of electron microscopy.The issue contains key papers by leading international researchers on the recent developments and applications of electron microscopy in the solid state and liquid state sciences. They include synthesis and characterization of silicon nitride nanorods, nanostructures of amorphous silica, electron microscopy studies of nanoscale structure and chemistry of Pt-Ru electrocatalysts of interest in direct methanol fuel cells, development of in situ wet-environmental transmission electron microscopy for the first nanoscale studies of dynamic liquid-catalyst reactions, strain analysis of silicon by finite element method and energy filtering convergent beam electron diffraction, applications of chemistry with electron microscopy, bismuth nanowires for applications in nanoelectronics technology, synthesis and characterization of quantum dots for superlattices and in situ electron microscopy at very high temperatures to study the motion of W5Si3 on [alpha][beta]-SiN3 substrates.We thank all the participants, including the invited speakers, contributors, and session chairs, who made the symposium successful. We also thank the authors and reviewers of the papers who worked assiduously towards the publication of this issue.We are very grateful to the Microscopy Society of America (MSA) for providing the opportunity to honor Professor Sir John Meurig Thomas. Organizational support from the MSA is also gratefully acknowledged.We thank Charles E. Lyman, editor in chief of Microscopy and Microanalysis for coordinating the publication of this issue and the entire journal staff for their efforts.

Local probe studies on lattice distortions and electronic correlations in manganites

NASA Astrophysics Data System (ADS)

Lopes, Armandina Maria Lima

Nesta tese apresenta-se um estudo experimental das distorcoes locais e correlacoes electronicas em oxidos magneticos com magnetoresistencia colossal. A tecnica de sonda local - Correlacao Angular Perturbada - e utilizada em amostras caracterizadas quanto as suas propriedades macroscopicas nomeadamente propriedades estruturais, magneticas e electricas, tendo em vista a obtencao de informacao microscopica relevante via gradiente de campo electrico e campo magnetico hiperfino, focando em particular os seguintes aspectos: -Distorcoes de rede e agregados de polaroes no sistema LaMnO3+Delta. E apresentado um estudo dos gradientes de campo electrica e campo magnetico hiperfino em amostras representativas do sistema LaMnO3+Delta, correlacionando estas propriedades locais com a caracterizacao das propriedades macroscopicas, efectuada nas mesmas amostras. Desta forma, foi possivel estudar a natureza microscopica das distorcoes polaronicas. Foi dada especial atencao ao composto com composicao LaMnO3.12 uma vez que este e um sistema padrao de uma manganite ferromagnetica-isoladora que apresenta uma transicao estrutural romboedrica (R)-ortorrombica (O) perto da temperatura ambiente. O estudo revelou que agregados de distorcoes locais sobrevivem ate 776 K, na fase de estrutura media mais simetrica (romboedrica), onde, por simetria, os octaedros MnO6 deveriam ser regulares. Estas distorcoes sao semelhantes as observadas no sistema LaMnO3 onde os octaedros MnO6 apresentam uma distorcao Jahn-Teller colectiva. Com a diminuicao da temperatura observa-se um aumento continuo destes agregados. Abaixo de uma temperatura critica estas distorcoes relaxam acomodando-se numa estrutura com reduzidas distorcoes Jahn-teller. Verificou-se tambem que a transicao estrutural (macroscopica) pode ser entendida como uma transicao de percolacao dos ambientes microscopicos. -Coexistencia das ordens electrica e magnetica no sistema Pr1-xCaxMnO3. E apresentado o primeiro estudo de gradiente de campo electrico no sistema Pr1-xCaxMnO3. Este estudo foi efectuado numa larga gama de temperaturas permitindo estudar localmente as diversas transicoes que este sistema apresenta. Em particular, na regiao do diagrama de fases onde existe ordenamento de carga e orbital (0.32

Tendências de teses e dissertações sobre ensino de astronomia no Brasil

NASA Astrophysics Data System (ADS)

Bretones, P. S.; Megid Neto, J.

2003-08-01

Neste trabalho são apresentados os resultados de uma pesquisa do tipo estado da arte sobre teses e dissertações defendidas no Brasil e relativas ao ensino de Astronomia. Teve por objetivo identificar essa produção e conhecer as principais tendências da pesquisa nesse campo. O procedimento inicial consistiu de um levantamento bibliográfico junto ao Centro de Documentação em Ensino de Ciências (CEDOC) da Faculdade de Educação da UNICAMP e ao Banco de Teses da CAPES disponível na Internet. Foram localizadas 13 dissertações de mestrado e 3 teses de doutorado, as quais foram estudadas em função dos seguintes aspectos: instituição, ano de defesa, nível escolar abrangido no estudo, foco temático do estudo e gênero de trabalho acadêmico. Deste conjunto de pesquisas, 13 (81,3%) delas foram defendidas a partir da segunda metade dos anos 90, indicando uma preocupação mais recente com temas relativos ao ensino de Astronomia no conjunto da produção acadêmica em programas de pós-graduação no Brasil. Verificou-se que 43,7% dos trabalhos foram produzidas na USP e 18,8% na UNICAMP. Quanto ao nível escolar abrangido nos estudos, predominaram os estudos direcionados ao Ensino Fundamental de 5a a 8a séries (62,5%). No que diz respeito ao foco temático das pesquisas, as principais tendências voltaram-se: 56,3% para Conteúdo e Método; 43,8% para Concepções do Professor; 37,5% para Currículo e Programas; 37,5% para Recursos Didáticos. Quanto ao gênero de trabalho acadêmico, verificou-se que 43,8% são de Pesquisa Experimental e 31,3% de Pesquisa de Análise de Conteúdo. Estudos de revisão bibliográfica como este visam colaborar com a divulgação ampla da produção acadêmica em determinada área, traçando algumas de suas tendências. Ao mesmo tempo possibilita, a partir de investigações decorrentes, apontar as suas contribuições para o ensino e sinalizar com necessidades a serem supridas por futuras pesquisas.

Soil Analysis Micro-Mission Concepts Derived from the MSP 2001 Mars Environmental Compatibility Assessment (MECA)

NASA Technical Reports Server (NTRS)

Hecht, M. H.; Meloy, T. P.; Anderson, M. S.; Buehler, M. G.; Frant, M. A.; Grannan, S. M.; Fuerstenau, S. D.; Keller, H. U.; Markiewicz, W. J.; Marshall, J.

1999-01-01

The Mars Environmental Compatibility Assessment (MECA) will evaluate the Martian environment for soil and dust-related hazards to human exploration as part of the Mars Surveyor Program 2001 Lander. The integrated MECA payload contains a wet-chemistry laboratory, a microscopy station, an electrometer to characterize the electrostatic environment, and arrays of material patches to study abrasion and adhesion. Heritage will be all-important for low cost micro-missions, and adaptations of instruments developed for the Pathfinder, '98 and '01 Landers should be strong contenders for '03 flights. This talk has three objectives: (1) Familiarize the audience with MECA instrument capabilities; (2) present concepts for stand-alone and/or mobile versions of MECA instruments; and (3) broaden the context of the MECA instruments from human exploration to a comprehensive scientific survey of Mars. Due to time limitations, emphasis will be on the chemistry and microscopy experiments. Ion-selective electrodes and related sensors in MECA's wet-chemistry laboratory will evaluate total dissolved solids, redox potential, pH, and the concentration of many soluble ions and gases in wet Martian soil. These electrodes can detect potentially dangerous heavy-metal ions, emitted pathogenic gases, and the soil's corrosive potential, and experiments will include cyclic voltammetry and anodic stripping. For experiments beyond 2001, enhancements could allow multiple use of the cells (for mobile experiments) and reagent addition (for quantitative mineralogical and exobiological analysis). MECA's microscopy station combines optical and atomic-force microscopy (AFM) in an actively focused, controlled illumination environment to image particles from millimeters to nanometers in size. Careful selection of substrates allows controlled experiments in adhesion, abrasion, hardness, aggregation, magnetic and other properties. Special tools allow primitive manipulation (brushing and scraping) of samples. Soil particle properties including size, shape, color, hardness, adhesive potential (electrostatic and magnetic), will be determined using an array of sample receptacles and collection substrates. The simple, rugged atomic-force microscope will image in the submicron size range and has the capability of performing a particle-by-particle analysis of the dust and soil. Future implementations might enhance the optical microscopy with spectroscopy, or incorporate advanced AFM techniques for thermogravimetric and chemical analysis.

Helium ion microscopy and ultra-high-resolution scanning electron microscopy analysis of membrane-extracted cells reveals novel characteristics of the cytoskeleton of Giardia intestinalis.

PubMed

Gadelha, Ana Paula Rocha; Benchimol, Marlene; de Souza, Wanderley

2015-06-01

Giardia intestinalis presents a complex microtubular cytoskeleton formed by specialized structures, such as the adhesive disk, four pairs of flagella, the funis and the median body. The ultrastructural organization of the Giardia cytoskeleton has been analyzed using different microscopic techniques, including high-resolution scanning electron microscopy. Recent advances in scanning microscopy technology have opened a new venue for the characterization of cellular structures and include scanning probe microscopy techniques such as ultra-high-resolution scanning electron microscopy (UHRSEM) and helium ion microscopy (HIM). Here, we studied the organization of the cytoskeleton of G. intestinalis trophozoites using UHRSEM and HIM in membrane-extracted cells. The results revealed a number of new cytoskeletal elements associated with the lateral crest and the dorsal surface of the parasite. The fine structure of the banded collar was also observed. The marginal plates were seen linked to a network of filaments, which were continuous with filaments parallel to the main cell axis. Cytoplasmic filaments that supported the internal structures were seen by the first time. Using anti-actin antibody, we observed a labeling in these filamentous structures. Taken together, these data revealed new surface characteristics of the cytoskeleton of G. intestinalis and may contribute to an improved understanding of the structural organization of trophozoites. Copyright © 2015 Elsevier Inc. All rights reserved.

Neurosurgical confocal endomicroscopy: A review of contrast agents, confocal systems, and future imaging modalities

PubMed Central

Zehri, Aqib H.; Ramey, Wyatt; Georges, Joseph F.; Mooney, Michael A.; Martirosyan, Nikolay L.; Preul, Mark C.; Nakaji, Peter

2014-01-01

Background: The clinical application of fluorescent contrast agents (fluorescein, indocyanine green, and aminolevulinic acid) with intraoperative microscopy has led to advances in intraoperative brain tumor imaging. Their properties, mechanism of action, history of use, and safety are analyzed in this report along with a review of current laser scanning confocal endomicroscopy systems. Additional imaging modalities with potential neurosurgical utility are also analyzed. Methods: A comprehensive literature search was performed utilizing PubMed and key words: In vivo confocal microscopy, confocal endomicroscopy, fluorescence imaging, in vivo diagnostics/neoplasm, in vivo molecular imaging, and optical imaging. Articles were reviewed that discussed clinically available fluorophores in neurosurgery, confocal endomicroscopy instrumentation, confocal microscopy systems, and intraoperative cancer diagnostics. Results: Current clinically available fluorescent contrast agents have specific properties that provide microscopic delineation of tumors when imaged with laser scanning confocal endomicroscopes. Other imaging modalities such as coherent anti-Stokes Raman scattering (CARS) microscopy, confocal reflectance microscopy, fluorescent lifetime imaging (FLIM), two-photon microscopy, and second harmonic generation may also have potential in neurosurgical applications. Conclusion: In addition to guiding tumor resection, intraoperative fluorescence and microscopy have the potential to facilitate tumor identification and complement frozen section analysis during surgery by providing real-time histological assessment. Further research, including clinical trials, is necessary to test the efficacy of fluorescent contrast agents and optical imaging instrumentation in order to establish their role in neurosurgery. PMID:24872922

Comparative Analysis Reveals Potential Utility of Digital Microscopy in the Evaluation of Peripheral Blood Smears With Some Barriers to Implementation.

PubMed

Gomez-Gelvez, Juan C; Kryvenko, Oleksandr N; Chabot-Richards, Devon S; Foucar, Kathryn; Inamdar, Kedar V; Karner, Kristin H

2015-07-01

Evaluation of the peripheral blood smear (PBS) is an essential diagnostic test in current medical practice. We aimed to evaluate the use of digital microscopy for the examination of PBS as an option to provide expert interpretation to remote sites and in "on-call" situations. We collected 100 Wright-Giemsa-stained PBS slides representing normal and abnormal findings seen at a community-based hospital. Four hematopathologists independently evaluated the cases using conventional light and digital microscopy. When comparing digital vs light microscopy, most of the cellular features evaluated showed at least a moderate degree of agreement in at least three of the reviewers. Discrepancies in final diagnosis were identified in a minority of the cases, most of which were attributed to the poorer resolution of digital microscopy at high magnification (×400). These results support the limited use of digital microscopy for evaluation and triage of peripheral blood smears as a practical option to obtain expert opinion in locations where experienced staff is not available on site. Our results indicate that while digital microscopy is well suited for basic triage of these blood smears, limitations in quality of imaging at higher magnification as well as large file size may limit its utility in certain settings and situations. Copyright© by the American Society for Clinical Pathology.

Green synthesis of silver nanoparticles using tannins

NASA Astrophysics Data System (ADS)

Raja, Pandian Bothi; Rahim, Afidah Abdul; Qureshi, Ahmad Kaleem; Awang, Khalijah

2014-09-01

Colloidal silver nanoparticles were prepared by rapid green synthesis using different tannin sources as reducing agent viz. chestnut (CN), mangrove (MG) and quebracho (QB). The aqueous silver ions when exposed to CN, MG and QB tannins were reduced which resulted in formation of silver nanoparticles. The resultant silver nanoparticles were characterized using UV-Visible, X-ray diffraction (XRD), scanning electron microscopy (SEM/EDX), and transmission electron microscopy (TEM) techniques. Furthermore, the possible mechanism of nanoparticles synthesis was also derived using FT-IR analysis. Spectroscopy analysis revealed that the synthesized nanoparticles were within 30 to 75 nm in size, while XRD results showed that nanoparticles formed were crystalline with face centered cubic geometry.

Multimodal biophotonic workstation for live cell analysis.

PubMed

Esseling, Michael; Kemper, Björn; Antkowiak, Maciej; Stevenson, David J; Chaudet, Lionel; Neil, Mark A A; French, Paul W; von Bally, Gert; Dholakia, Kishan; Denz, Cornelia

2012-01-01

A reliable description and quantification of the complex physiology and reactions of living cells requires a multimodal analysis with various measurement techniques. We have investigated the integration of different techniques into a biophotonic workstation that can provide biological researchers with these capabilities. The combination of a micromanipulation tool with three different imaging principles is accomplished in a single inverted microscope which makes the results from all the techniques directly comparable. Chinese Hamster Ovary (CHO) cells were manipulated by optical tweezers while the feedback was directly analyzed by fluorescence lifetime imaging, digital holographic microscopy and dynamic phase-contrast microscopy. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The measurement of hemoglobin oxygen saturation using multiwavelength photoacoustic microscopy

NASA Astrophysics Data System (ADS)

Deng, Zilin; Yang, Xiaoquan; Yu, Lejun; Gong, Hui

2010-02-01

Hemoglobin oxygen saturation (SO2) is one of the most critical functional parameters to the metabolism. In this paper, we mainly introduced some initial results of measuring blood oxygen using multi-wavelength photoacoustic microscopy (PAM). In phantom study, we demonstrate the photoacoustic signal amplitude increases linearly with the concentration of red or blue ink. Then the calculated concentration of red ink in double-ink mixtures with PAM has a 5% difference with the result measured with spectrophotometric analysis. In ex vivo experiment, the measured result exhibt 15% difference between the PAM and spectrophotometric analysis. Experiment results suggest that PAM could be used to determine the SO2 quantitatively.

The use of analytical surface tools in the fundamental study of wear. [atomic nature of wear

NASA Technical Reports Server (NTRS)

Buckley, D. H.

1977-01-01

Various techniques and surface tools available for the study of the atomic nature of the wear of materials are reviewed These include chemical etching, x-ray diffraction, electron diffraction, scanning electron microscopy, low-energy electron diffraction, Auger emission spectroscopy analysis, electron spectroscopy for chemical analysis, field ion microscopy, and the atom probe. Properties of the surface and wear surface regions which affect wear, such as surface energy, crystal structure, crystallographic orientation, mode of dislocation behavior, and cohesive binding, are discussed. A number of mechanisms involved in the generation of wear particles are identified with the aid of the aforementioned tools.

Failure Surface Analysis of Polyimide/Titanium Notched Coating Adhesion Specimens

DOE Office of Scientific and Technical Information (OSTI.GOV)

GIUNTA,RACHEL K.; KANDER,RONALD G.

2000-12-18

Adhesively bonded joints of LaRC{trademark} PETI-5, a phenylethynyl-terminated polyimide, with chromic acid anodized titanium were fabricated and debonded interfacially. The adhesive-substrate failure surfaces were investigated using several surface analysis techniques. From Auger spectroscopy, field emission scanning electron microscopy, and atomic force microscopy studies, polymer appears to be penetrating the pores of the anodized substrate to a depth of approximately 100 nm. From x-ray photoelectron spectroscopy data, the polymer penetrating the pores appears to be in electrical contact with the titanium substrate, leading to differential charging. These analyses confirm that the polymer is becoming mechanically interlocked within the substrate surface.

Stress corrosion in titanium alloys and other metallic materials

NASA Technical Reports Server (NTRS)

Harkins, C. G. (Editor); Brotzen, F. R.; Hightower, J. W.; Mclellan, R. B.; Roberts, J. M.; Rudee, M. L.; Leith, I. R.; Basu, P. K.; Salama, K.; Parris, D. P.

1971-01-01

Multiple physical and chemical techniques including mass spectroscopy, atomic absorption spectroscopy, gas chromatography, electron microscopy, optical microscopy, electronic spectroscopy for chemical analysis (ESCA), infrared spectroscopy, nuclear magnetic resonance (NMR), X-ray analysis, conductivity, and isotopic labeling were used in investigating the atomic interactions between organic environments and titanium and titanium oxide surfaces. Key anhydrous environments studied included alcohols, which contain hydrogen; carbon tetrachloride, which does not contain hydrogen; and mixtures of alcohols and halocarbons. Effects of dissolved salts in alcohols were also studied. This program emphasized experiments designed to delineate the conditions necessary rather than sufficient for initiation processes and for propagation processes in Ti SCC.

Green synthesis of multifunctional carbon dots from coriander leaves and their potential application as antioxidants, sensors and bioimaging agents.

PubMed

Sachdev, Abhay; Gopinath, P

2015-06-21

In the present study, a facile one-step hydrothermal treatment of coriander leaves for preparing carbon dots (CDs) has been reported. Optical and structural properties of the CDs have been extensively studied by UV-visible and fluorescence spectroscopic, microscopic (transmission electron microscopy, scanning electron microscopy) and X-ray diffraction techniques. Surface functionality and composition of the CDs have been illustrated by elemental analysis and Fourier transform infrared spectroscopy (FTIR). Quenching of the fluorescence of the CDs in the presence of metal ions is of prime significance, hence CDs have been used as a fluorescence probe for sensitive and selective detection of Fe(3+) ions. Eventually, biocompatibility and bioimaging aspects of CDs have been evaluated on lung normal (L-132) and cancer (A549) cell lines. Qualitative analysis of cellular uptake of CDs has been pursued through fluorescence microscopy, while quantitative analysis using a flow cytometer provided an insight into the concentration and cell-type dependent uptake of CDs. The article further investigates the antioxidant activity of CDs. Therefore, we have validated the practicality of CDs obtained from a herbal carbon source for versatile applications.

microMS: A Python Platform for Image-Guided Mass Spectrometry Profiling

NASA Astrophysics Data System (ADS)

Comi, Troy J.; Neumann, Elizabeth K.; Do, Thanh D.; Sweedler, Jonathan V.

2017-09-01

Image-guided mass spectrometry (MS) profiling provides a facile framework for analyzing samples ranging from single cells to tissue sections. The fundamental workflow utilizes a whole-slide microscopy image to select targets of interest, determine their spatial locations, and subsequently perform MS analysis at those locations. Improving upon prior reported methodology, a software package was developed for working with microscopy images. microMS, for microscopy-guided mass spectrometry, allows the user to select and profile diverse samples using a variety of target patterns and mass analyzers. Written in Python, the program provides an intuitive graphical user interface to simplify image-guided MS for novice users. The class hierarchy of instrument interactions permits integration of new MS systems while retaining the feature-rich image analysis framework. microMS is a versatile platform for performing targeted profiling experiments using a series of mass spectrometers. The flexibility in mass analyzers greatly simplifies serial analyses of the same targets by different instruments. The current capabilities of microMS are presented, and its application for off-line analysis of single cells on three distinct instruments is demonstrated. The software has been made freely available for research purposes. [Figure not available: see fulltext.

microMS: A Python Platform for Image-Guided Mass Spectrometry Profiling.

PubMed

Comi, Troy J; Neumann, Elizabeth K; Do, Thanh D; Sweedler, Jonathan V

2017-09-01

Image-guided mass spectrometry (MS) profiling provides a facile framework for analyzing samples ranging from single cells to tissue sections. The fundamental workflow utilizes a whole-slide microscopy image to select targets of interest, determine their spatial locations, and subsequently perform MS analysis at those locations. Improving upon prior reported methodology, a software package was developed for working with microscopy images. microMS, for microscopy-guided mass spectrometry, allows the user to select and profile diverse samples using a variety of target patterns and mass analyzers. Written in Python, the program provides an intuitive graphical user interface to simplify image-guided MS for novice users. The class hierarchy of instrument interactions permits integration of new MS systems while retaining the feature-rich image analysis framework. microMS is a versatile platform for performing targeted profiling experiments using a series of mass spectrometers. The flexibility in mass analyzers greatly simplifies serial analyses of the same targets by different instruments. The current capabilities of microMS are presented, and its application for off-line analysis of single cells on three distinct instruments is demonstrated. The software has been made freely available for research purposes. Graphical Abstract ᅟ.

A high-level 3D visualization API for Java and ImageJ.

PubMed

Schmid, Benjamin; Schindelin, Johannes; Cardona, Albert; Longair, Mark; Heisenberg, Martin

2010-05-21

Current imaging methods such as Magnetic Resonance Imaging (MRI), Confocal microscopy, Electron Microscopy (EM) or Selective Plane Illumination Microscopy (SPIM) yield three-dimensional (3D) data sets in need of appropriate computational methods for their analysis. The reconstruction, segmentation and registration are best approached from the 3D representation of the data set. Here we present a platform-independent framework based on Java and Java 3D for accelerated rendering of biological images. Our framework is seamlessly integrated into ImageJ, a free image processing package with a vast collection of community-developed biological image analysis tools. Our framework enriches the ImageJ software libraries with methods that greatly reduce the complexity of developing image analysis tools in an interactive 3D visualization environment. In particular, we provide high-level access to volume rendering, volume editing, surface extraction, and image annotation. The ability to rely on a library that removes the low-level details enables concentrating software development efforts on the algorithm implementation parts. Our framework enables biomedical image software development to be built with 3D visualization capabilities with very little effort. We offer the source code and convenient binary packages along with extensive documentation at http://3dviewer.neurofly.de.

Investigating the Moisture Content of Polyamide 6 by Raman-Microscopy and Multivariate Data Analysis

NASA Astrophysics Data System (ADS)

Lechner, Tobias; Noack, Kristina; Thöne, Manuel; Amend, Philipp; Schmidt, Michael; Will, Stefan

Thermal malleability of thermoplastics results in a high product diversity in various industry sectors. However, industrial applications require a constant and high component quality. Hence, material processing such as laser welding has to consider that, e.g., the moisture content of thermoplastics influences the mechanical properties such as the tensile strength. Moreover, water evaporates during laser welding and can form pores and defects. Thus, there is a large need for non-invasive material inspection before processing. To that end, we developed a methodology based on Raman-microscopy and multivariate data analysis (MVD) to determine the moisture content of polyamide (MCP). Further, the impact of the MCP on the mechanical properties was verified. For samples with a defined variation of the MCP, xyz-Raman-scans were carried out and analysed using MVD. For reference purposes, the samples were weighted and tensile tests were performed. An evaluation by means of partial least squares regression analysis (PLSR) resulted in a prediction of the MCP with a correlation coefficient >98%. Consequently, Raman-microscopy shows large potential for developing new techniques for inspection and quality control of plastics before processing. Dedicated to Professor Alfred Leipertz on the occasion of his 70th birthday.

Scanning transmission ion microscopy mass measurements for quantitative trace element analysis within biological samples and validation using atomic force microscopy thickness measurements

NASA Astrophysics Data System (ADS)

Devès, Guillaume; Cohen-Bouhacina, Touria; Ortega, Richard

2004-10-01

We used the nuclear microprobe techniques, micro-PIXE (particle-induced X-ray emission), micro-RBS (Rutherford backscattering spectrometry) and scanning transmission ion microscopy (STIM) in order to perform the characterization of trace element content and spatial distribution within biological samples (dehydrated cultured cells, tissues). The normalization of PIXE results was usually expressed in terms of sample dry mass as determined by micro-RBS recorded simultaneously to micro-PIXE. However, the main limit of RBS mass measurement is the sample mass loss occurring during irradiation and which could be up to 30% of the initial sample mass. We present here a new methodology for PIXE normalization and quantitative analysis of trace element within biological samples based on dry mass measurement performed by mean of STIM. The validation of STIM cell mass measurements was obtained in comparison with AFM sample thickness measurements. Results indicated the reliability of STIM mass measurement performed on biological samples and suggested that STIM should be performed for PIXE normalization. Further information deriving from direct confrontation of AFM and STIM analysis could as well be obtained, like in situ measurements of cell specific gravity within cells compartment (nucleolus and cytoplasm).

Scanning electron microscope fractography in failure analysis of steels

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wouters, R.; Froyen, L.

1996-04-01

For many failure cases, macroscopic examination of the fracture surface permits discrimination of fatigue fractures from overload fractures. For clarifying fatigue fractures, the practical significance of microfractography is limited to an investigation of the crack initiation areas. Scanning electron microscopy is successfully used in tracing local material abnormalities that act as fatigue crack initiators. The task for the scanning electron microscope, however, is much more substantial in failure analysis of overload fractures, especially for steels. By revealing specific fractographic characteristics, complemented by information about the material and the loading conditions, scanning electron microscopy provides a strong indication of the probablemore » cause of failure. A complete dimple fracture is indicative of acceptable bulk material properties; overloading, by subdimensioning or excessive external loading, has to be verified. The presence of cleavage fracture makes the material properties questionable if external conditions causing embrittlement are absent. Intergranular brittle fracture requires verification of grain-boundary weakening conditions--a sensitized structure, whether or not combined with a local stress state or a specific environment. The role of scanning electron microscopy in failure analysis is illustrated by case histories of the aforementioned fracture types.« less

Brain tumor classification using AFM in combination with data mining techniques.

PubMed

Huml, Marlene; Silye, René; Zauner, Gerald; Hutterer, Stephan; Schilcher, Kurt

2013-01-01

Although classification of astrocytic tumors is standardized by the WHO grading system, which is mainly based on microscopy-derived, histomorphological features, there is great interobserver variability. The main causes are thought to be the complexity of morphological details varying from tumor to tumor and from patient to patient, variations in the technical histopathological procedures like staining protocols, and finally the individual experience of the diagnosing pathologist. Thus, to raise astrocytoma grading to a more objective standard, this paper proposes a methodology based on atomic force microscopy (AFM) derived images made from histopathological samples in combination with data mining techniques. By comparing AFM images with corresponding light microscopy images of the same area, the progressive formation of cavities due to cell necrosis was identified as a typical morphological marker for a computer-assisted analysis. Using genetic programming as a tool for feature analysis, a best model was created that achieved 94.74% classification accuracy in distinguishing grade II tumors from grade IV ones. While utilizing modern image analysis techniques, AFM may become an important tool in astrocytic tumor diagnosis. By this way patients suffering from grade II tumors are identified unambiguously, having a less risk for malignant transformation. They would benefit from early adjuvant therapies.

Investigation of Microstructure and Mechanical Properties of ECAP-Processed AM Series Magnesium Alloy

NASA Astrophysics Data System (ADS)

Gopi, K. R.; Nayaka, H. Shivananda; Sahu, Sandeep

2016-09-01

Magnesium alloy Mg-Al-Mn (AM70) was processed by equal channel angular pressing (ECAP) at 275 °C for up to 4 passes in order to produce ultrafine-grained microstructure and improve its mechanical properties. ECAP-processed samples were characterized for microstructural analysis using optical microscopy, scanning electron microscopy, and transmission electron microscopy. Microstructural analysis showed that, with an increase in the number of ECAP passes, grains refined and grain size reduced from an average of 45 to 1 µm. Electron backscatter diffraction analysis showed the transition from low angle grain boundaries to high angle grain boundaries in ECAP 4 pass sample as compared to as-cast sample. The strength and hardness values an showed increasing trend for the initial 2 passes of ECAP processing and then started decreasing with further increase in the number of ECAP passes, even though the grain size continued to decrease in all the successive ECAP passes. However, the strength and hardness values still remained quite high when compared to the initial condition. This behavior was found to be correlated with texture modification in the material as a result of ECAP processing.

Reactive Brazing of Carbon-Carbon Composites to Titanium

NASA Technical Reports Server (NTRS)

Shpargel, Tarah; Singh, M.; Morscher, Gregory; Asthana, Rajiv

2004-01-01

The Ti-metal/C-C composite joints were formed by reactive brazing with three commercial brazes, namely, Cu-ABA, TiCuNi, and TiCuSil. The joint microstructures were examined using optical microscopy, and scanning electron microscopy (SEM) coupled with energy dispersive spectrometry (EDS). The results of the microstructure analysis indicate solute redistribution across the joint and possible metallurgical bond formation via interdiffusion, which led to good wetting and spreading.

Facile synthesis of silicon nanowire-nanopillar superhydrophobic structures

NASA Astrophysics Data System (ADS)

Roy, Abhijit; Satpati, Biswarup

2018-04-01

We have used metal assisted chemical etching (MACE) method to produce silicon (Si) nanowire-nanopillar array. Nanowire-nanopillar combined structures show higher degree of hydrophobicity compared to its nanowire (Si-NW) counterparts. The rate of etching is depended on initial metal deposition. The structural analysis was carried out using scanning electron microscopy (SEM) in combination with transmission electron microscopy (TEM) to determine different parameters like etching direction, crystallinity etc.

Active Metal Brazing of Carbon-Carbon Composites to Titanium

NASA Technical Reports Server (NTRS)

Singh, M.; Shpargel, T. P.; Morscher, G.; Asthana, R.

2004-01-01

The Ti-metal/C-C composite joints were formed by reactive brazing with three commercial brazes, namely, Cu-ABA, TiCuNi, and TiCuSil. The joint microstructures were examined using optical microscopy, and scanning electron microscopy (SEM) coupled with energy dispersive spectrometry (EDS). The results of the microstructure analysis indicate solute redistribution across the joint which led to good wetting, spreading, and metallurgical bond formation via interdiffusion.

Microelectronics Failure Analysis Techniques. A Procedural Guide

DTIC Science & Technology

1980-01-01

Process Control ." Scanning Electron Microscopy/1976, lIT Research Institute, Chicago, Illinois, April 1976, pp. 515-519. 49. Cunningham, R.F. "Sample...Character- ization and Quality Control ." Scanning Electron Microscopy/1977 vol. I, lIT Research Institute, Chicago, Illinois, March 1977, pp. 201-210. 151...reference purposes. Neither the United States Government, the General Electric Company nor the lIT Research Institute warrant the accuracy of this

Application of Electron Microscopy Techniques to the Investigation of Space Shuttle Columbia Accident

NASA Technical Reports Server (NTRS)

Shah, Sandeep

2005-01-01

This viewgraph presentation gives an overview of the investigation into the breakup of the Space Shuttle Columbia, and addresses the importance of a failure analysis strategy for the investigation of the Columbia accident. The main focus of the presentation is on the usefulness of electron microscopy for analyzing slag deposits from the tiles and reinforced carbon-carbon (RCC) wing panels of the Columbia orbiter.

Dynamic longitudinal investigation of individual nerve endings in the skin of anesthetized mice using in vivo two-photon microscopy

NASA Astrophysics Data System (ADS)

Yuryev, Mikhail; Khiroug, Leonard

2012-04-01

Visualization of individual cutaneous nerve endings has previously relied on laborious procedures of tissue excision, fixation, sectioning and staining for light or electron microscopy. We present a method for non-invasive, longitudinal two-photon microscopy of single nerve endings within the skin of anesthetized transgenic mice. Besides excellent signal-to-background ratio and nanometer-scale spatial resolution, this method offers time-lapse ``movies'' of pathophysiological changes in nerve fine structure over minutes, hours, days or weeks. Structure of keratinocytes and dermal matrix is visualized simultaneously with nerve endings, providing clear landmarks for longitudinal analysis. We further demonstrate feasibility of dissecting individual nerve fibers with infra-red laser and monitoring their degradation and regeneration. In summary, our excision-free optical biopsy technique is ideal for longitudinal microscopic analysis of animal skin and skin innervations in vivo and can be applied widely in preclinical models of chronic pain, allergies, skin cancers and a variety of dermatological disorders.

SNSMIL, a real-time single molecule identification and localization algorithm for super-resolution fluorescence microscopy

PubMed Central

Tang, Yunqing; Dai, Luru; Zhang, Xiaoming; Li, Junbai; Hendriks, Johnny; Fan, Xiaoming; Gruteser, Nadine; Meisenberg, Annika; Baumann, Arnd; Katranidis, Alexandros; Gensch, Thomas

2015-01-01

Single molecule localization based super-resolution fluorescence microscopy offers significantly higher spatial resolution than predicted by Abbe’s resolution limit for far field optical microscopy. Such super-resolution images are reconstructed from wide-field or total internal reflection single molecule fluorescence recordings. Discrimination between emission of single fluorescent molecules and background noise fluctuations remains a great challenge in current data analysis. Here we present a real-time, and robust single molecule identification and localization algorithm, SNSMIL (Shot Noise based Single Molecule Identification and Localization). This algorithm is based on the intrinsic nature of noise, i.e., its Poisson or shot noise characteristics and a new identification criterion, QSNSMIL, is defined. SNSMIL improves the identification accuracy of single fluorescent molecules in experimental or simulated datasets with high and inhomogeneous background. The implementation of SNSMIL relies on a graphics processing unit (GPU), making real-time analysis feasible as shown for real experimental and simulated datasets. PMID:26098742

High-resolution measurements of the multilayer ultra-structure of articular cartilage and their translational potential

PubMed Central

2014-01-01

Current musculoskeletal imaging techniques usually target the macro-morphology of articular cartilage or use histological analysis. These techniques are able to reveal advanced osteoarthritic changes in articular cartilage but fail to give detailed information to distinguish early osteoarthritis from healthy cartilage, and this necessitates high-resolution imaging techniques measuring cells and the extracellular matrix within the multilayer structure of articular cartilage. This review provides a comprehensive exploration of the cellular components and extracellular matrix of articular cartilage as well as high-resolution imaging techniques, including magnetic resonance image, electron microscopy, confocal laser scanning microscopy, second harmonic generation microscopy, and laser scanning confocal arthroscopy, in the measurement of multilayer ultra-structures of articular cartilage. This review also provides an overview for micro-structural analysis of the main components of normal or osteoarthritic cartilage and discusses the potential and challenges associated with developing non-invasive high-resolution imaging techniques for both research and clinical diagnosis of early to late osteoarthritis. PMID:24946278

High throughput, detailed, cell-specific neuroanatomy of dendritic spines using microinjection and confocal microscopy

PubMed Central

Dumitriu, Dani; Rodriguez, Alfredo; Morrison, John H.

2012-01-01

Morphological features such as size, shape and density of dendritic spines have been shown to reflect important synaptic functional attributes and potential for plasticity. Here we describe in detail a protocol for obtaining detailed morphometric analysis of spines using microinjection of fluorescent dyes, high resolution confocal microscopy, deconvolution and image analysis using NeuronStudio. Recent technical advancements include better preservation of tissue resulting in prolonged ability to microinject, and algorithmic improvements that compensate for the residual Z-smear inherent in all optical imaging. Confocal imaging parameters were probed systematically for the identification of both optimal resolution as well as highest efficiency. When combined, our methods yield size and density measurements comparable to serial section transmission electron microscopy in a fraction of the time. An experiment containing 3 experimental groups with 8 subjects in each can take as little as one month if optimized for speed, or approximately 4 to 5 months if the highest resolution and morphometric detail is sought. PMID:21886104

Image Quality Ranking Method for Microscopy

PubMed Central

Koho, Sami; Fazeli, Elnaz; Eriksson, John E.; Hänninen, Pekka E.

2016-01-01

Automated analysis of microscope images is necessitated by the increased need for high-resolution follow up of events in time. Manually finding the right images to be analyzed, or eliminated from data analysis are common day-to-day problems in microscopy research today, and the constantly growing size of image datasets does not help the matter. We propose a simple method and a software tool for sorting images within a dataset, according to their relative quality. We demonstrate the applicability of our method in finding good quality images in a STED microscope sample preparation optimization image dataset. The results are validated by comparisons to subjective opinion scores, as well as five state-of-the-art blind image quality assessment methods. We also show how our method can be applied to eliminate useless out-of-focus images in a High-Content-Screening experiment. We further evaluate the ability of our image quality ranking method to detect out-of-focus images, by extensive simulations, and by comparing its performance against previously published, well-established microscopy autofocus metrics. PMID:27364703

Phenol homeostasis is ensured in vanilla fruit by storage under solid form in a new chloroplast-derived organelle, the phenyloplast

PubMed Central

Conéjéro, Geneviève

2014-01-01

A multiple cell imaging approach combining immunofluorescence by confocal microscopy, fluorescence spectral analysis by multiphotonic microscopy, and transmission electron microscopy identified the site of accumulation of 4-O-(3-methoxybenzaldehyde) β-d-glucoside, a phenol glucoside massively stockpiled by vanilla fruit. The glucoside is sufficiently abundant to be detected by spectral analysis of its autofluorescence. The convergent results obtained by these different techniques demonstrated that the phenol glucoside accumulates in the inner volume of redifferentiating chloroplasts as solid amorphous deposits, thus ensuring phenylglucoside cell homeostasis. Redifferentiation starts with the generation of loculi between thylakoid membranes which are progressively filled with the glucoside until a fully matured organelle is obtained. This peculiar mode of storage of a phenolic secondary metabolite is suspected to occur in other plants and its generalization in the Plantae could be considered. This new chloroplast-derived organelle is referred to as a ‘phenyloplast’. PMID:24683183

Phenol homeostasis is ensured in vanilla fruit by storage under solid form in a new chloroplast-derived organelle, the phenyloplast.

PubMed

Brillouet, Jean-Marc; Verdeil, Jean-Luc; Odoux, Eric; Lartaud, Marc; Grisoni, Michel; Conéjéro, Geneviève

2014-06-01

A multiple cell imaging approach combining immunofluorescence by confocal microscopy, fluorescence spectral analysis by multiphotonic microscopy, and transmission electron microscopy identified the site of accumulation of 4-O-(3-methoxybenzaldehyde) β-d-glucoside, a phenol glucoside massively stockpiled by vanilla fruit. The glucoside is sufficiently abundant to be detected by spectral analysis of its autofluorescence. The convergent results obtained by these different techniques demonstrated that the phenol glucoside accumulates in the inner volume of redifferentiating chloroplasts as solid amorphous deposits, thus ensuring phenylglucoside cell homeostasis. Redifferentiation starts with the generation of loculi between thylakoid membranes which are progressively filled with the glucoside until a fully matured organelle is obtained. This peculiar mode of storage of a phenolic secondary metabolite is suspected to occur in other plants and its generalization in the Plantae could be considered. This new chloroplast-derived organelle is referred to as a 'phenyloplast'. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Novel microscopy-based screening method reveals regulators of contact-dependent intercellular transfer

PubMed Central

Michael Frei, Dominik; Hodneland, Erlend; Rios-Mondragon, Ivan; Burtey, Anne; Neumann, Beate; Bulkescher, Jutta; Schölermann, Julia; Pepperkok, Rainer; Gerdes, Hans-Hermann; Kögel, Tanja

2015-01-01

Contact-dependent intercellular transfer (codeIT) of cellular constituents can have functional consequences for recipient cells, such as enhanced survival and drug resistance. Pathogenic viruses, prions and bacteria can also utilize this mechanism to spread to adjacent cells and potentially evade immune detection. However, little is known about the molecular mechanism underlying this intercellular transfer process. Here, we present a novel microscopy-based screening method to identify regulators and cargo of codeIT. Single donor cells, carrying fluorescently labelled endocytic organelles or proteins, are co-cultured with excess acceptor cells. CodeIT is quantified by confocal microscopy and image analysis in 3D, preserving spatial information. An siRNA-based screening using this method revealed the involvement of several myosins and small GTPases as codeIT regulators. Our data indicates that cellular protrusions and tubular recycling endosomes are important for codeIT. We automated image acquisition and analysis to facilitate large-scale chemical and genetic screening efforts to identify key regulators of codeIT. PMID:26271723

Analysis of gene expression levels in individual bacterial cells without image segmentation.

PubMed

Kwak, In Hae; Son, Minjun; Hagen, Stephen J

2012-05-11

Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly. Copyright © 2012 Elsevier Inc. All rights reserved.

3D Filament Network Segmentation with Multiple Active Contours

NASA Astrophysics Data System (ADS)

Xu, Ting; Vavylonis, Dimitrios; Huang, Xiaolei

2014-03-01

Fluorescence microscopy is frequently used to study two and three dimensional network structures formed by cytoskeletal polymer fibers such as actin filaments and microtubules. While these cytoskeletal structures are often dilute enough to allow imaging of individual filaments or bundles of them, quantitative analysis of these images is challenging. To facilitate quantitative, reproducible and objective analysis of the image data, we developed a semi-automated method to extract actin networks and retrieve their topology in 3D. Our method uses multiple Stretching Open Active Contours (SOACs) that are automatically initialized at image intensity ridges and then evolve along the centerlines of filaments in the network. SOACs can merge, stop at junctions, and reconfigure with others to allow smooth crossing at junctions of filaments. The proposed approach is generally applicable to images of curvilinear networks with low SNR. We demonstrate its potential by extracting the centerlines of synthetic meshwork images, actin networks in 2D TIRF Microscopy images, and 3D actin cable meshworks of live fission yeast cells imaged by spinning disk confocal microscopy.

Precipitation and Phase Transformations in 2101 Lean Duplex Stainless Steel During Isothermal Aging

NASA Astrophysics Data System (ADS)

Maetz, Jean-Yves; Cazottes, Sophie; Verdu, Catherine; Kleber, Xavier

2016-01-01

The effect of isothermal aging at 963 K (690 °C) on the microstructure of a 2101 lean duplex stainless steel, with the composition Fe-21.5Cr-5Mn-1.6Ni-0.22N-0.3Mo, was investigated using a multi-technique and multi-scale approach. The kinetics of phase transformation and precipitation was followed from a few minutes to thousands of hours using thermoelectric power measurements; based on these results, certain aging states were selected for electron microscopy characterization. Scanning electron microscopy, electron back-scattered diffraction, and transmission electron microscopy were used to quantitatively describe the microstructural evolution through crystallographic analysis, chemical analysis, and volume fraction measurements from the macroscopic scale down to the nanometric scale. During aging, the precipitation of M23C6 carbides, Cr2N nitrides, and σ phase as well as the transformation of ferrite into austenite and austenite into martensite was observed. These complex microstructural changes are controlled by Cr volume diffusion. The precipitation and phase transformation mechanisms are described.

Digital holographic microscopy

NASA Astrophysics Data System (ADS)

Barkley, Solomon; Dimiduk, Thomas; Manoharan, Vinothan

Digital holographic microscopy is a 3D optical imaging technique with high temporal ( ms) and spatial ( 10 nm) precision. However, its adoption as a characterization technique has been limited due to the inherent difficulty of recovering 3D data from the holograms. Successful analysis has traditionally required substantial knowledge about the sample being imaged (for example, the approximate positions of particles in the field of view), as well as expertise in scattering theory. To overcome the obstacles to widespread adoption of holographic microscopy, we developed HoloPy - an open source python package for analysis of holograms and scattering data. HoloPy uses Bayesian statistical methods to determine the geometry and properties of discrete scatterers from raw holograms. We demonstrate the use of HoloPy to measure the dynamics of colloidal particles at interfaces, to ascertain the structures of self-assembled colloidal particles, and to track freely swimming bacteria. The HoloPy codebase is thoroughly tested and well-documented to facilitate use by the broader experimental community. This research is supported by NSF Grant DMR-1306410 and NSERC.

Measuring single-cell gene expression dynamics in bacteria using fluorescence time-lapse microscopy

PubMed Central

Young, Jonathan W; Locke, James C W; Altinok, Alphan; Rosenfeld, Nitzan; Bacarian, Tigran; Swain, Peter S; Mjolsness, Eric; Elowitz, Michael B

2014-01-01

Quantitative single-cell time-lapse microscopy is a powerful method for analyzing gene circuit dynamics and heterogeneous cell behavior. We describe the application of this method to imaging bacteria by using an automated microscopy system. This protocol has been used to analyze sporulation and competence differentiation in Bacillus subtilis, and to quantify gene regulation and its fluctuations in individual Escherichia coli cells. The protocol involves seeding and growing bacteria on small agarose pads and imaging the resulting microcolonies. Images are then reviewed and analyzed using our laboratory's custom MATLAB analysis code, which segments and tracks cells in a frame-to-frame method. This process yields quantitative expression data on cell lineages, which can illustrate dynamic expression profiles and facilitate mathematical models of gene circuits. With fast-growing bacteria, such as E. coli or B. subtilis, image acquisition can be completed in 1 d, with an additional 1–2 d for progressing through the analysis procedure. PMID:22179594

Molecular and ultrastructural analysis of forisome subunits reveals the principles of forisome assembly

PubMed Central

Müller, Boje; Groscurth, Sira; Menzel, Matthias; Rüping, Boris A.; Twyman, Richard M.; Prüfer, Dirk; Noll, Gundula A.

2014-01-01

Background and Aims Forisomes are specialized structural phloem proteins that mediate sieve element occlusion after wounding exclusively in papilionoid legumes, but most studies of forisome structure and function have focused on the Old World clade rather than the early lineages. A comprehensive phylogenetic, molecular, structural and functional analysis of forisomes from species covering a broad spectrum of the papilionoid legumes was therefore carried out, including the first analysis of Dipteryx panamensis forisomes, representing the earliest branch of the Papilionoideae lineage. The aim was to study the molecular, structural and functional conservation among forisomes from different tribes and to establish the roles of individual forisome subunits. Methods Sequence analysis and bioinformatics were combined with structural and functional analysis of native forisomes and artificial forisome-like protein bodies, the latter produced by expressing forisome genes from different legumes in a heterologous background. The structure of these bodies was analysed using a combination of confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and the function of individual subunits was examined by combinatorial expression, micromanipulation and light microscopy. Key Results Dipteryx panamensis native forisomes and homomeric protein bodies assembled from the single sieve element occlusion by forisome (SEO-F) subunit identified in this species were structurally and functionally similar to forisomes from the Old World clade. In contrast, homomeric protein bodies assembled from individual SEO-F subunits from Old World species yielded artificial forisomes differing in proportion to their native counterparts, suggesting that multiple SEO-F proteins are required for forisome assembly in these plants. Structural differences between Medicago truncatula native forisomes, homomeric protein bodies and heteromeric bodies containing all possible subunit combinations suggested that combinations of SEO-F proteins may fine-tune the geometric proportions and reactivity of forisomes. Conclusions It is concluded that forisome structure and function have been strongly conserved during evolution and that species-dependent subsets of SEO-F proteins may have evolved to fine-tune the structure of native forisomes. PMID:24694827

Conformational Switching in PolyGln Amyloid Fibrils Resulting from a Single Amino Acid Insertion

PubMed Central

Huang, Rick K.; Baxa, Ulrich; Aldrian, Gudrun; Ahmed, Abdullah B.; Wall, Joseph S.; Mizuno, Naoko; Antzutkin, Oleg; Steven, Alasdair C.; Kajava, Andrey V.

2014-01-01

The established correlation between neurodegenerative disorders and intracerebral deposition of polyglutamine aggregates motivates attempts to better understand their fibrillar structure. We designed polyglutamines with a few lysines inserted to overcome the hindrance of extreme insolubility and two D-lysines to limit the lengths of β-strands. One is 33 amino acids long (PolyQKd-33) and the other has one fewer glutamine (PolyQKd-32). Both form well-dispersed fibrils suitable for analysis by electron microscopy. Electron diffraction confirmed cross-β structures in both fibrils. Remarkably, the deletion of just one glutamine residue from the middle of the peptide leads to substantially different amyloid structures. PolyQKd-32 fibrils are consistently 10–20% wider than PolyQKd-33, as measured by negative staining, cryo-electron microscopy, and scanning transmission electron microscopy. Scanning transmission electron microscopy analysis revealed that the PolyQKd-32 fibrils have 50% higher mass-per-length than PolyQKd-33. This distinction can be explained by a superpleated β-structure model for PolyQKd-33 and a model with two β-solenoid protofibrils for PolyQKd-32. These data provide evidence for β-arch-containing structures in polyglutamine fibrils and open future possibilities for structure-based drug design. PMID:24853742

Web-based virtual microscopy at the RWTH Aachen University: didactic concept, methods and analysis of acceptance by the students.

PubMed

Merk, Magdalene; Knuechel, Ruth; Perez-Bouza, Alberto

2010-12-20

Fundamental knowledge of microscopic anatomy and pathology has always been an essential part in medical education. The traditional didactic concept comprises theoretical and practical lessons using a light microscope and glass slides. High-speed Internet connections and technical improvement in whole-slide digital microscopy (commonly termed "virtual microscopy") provide a new and attractive approach for both teachers and students. High picture quality and unlimited temporal and spatial availability of histology samples from different fields are key advantages of web-based digital microscopy. In this report we discuss the technical requirements, system efficiency, optical resolution and didactic concept. Furthermore, we present a review of the experience gained in the course of one year based on an analysis of student acceptance. Three groups with a total of 192 students between the 3rd and 5th year of medical studies attending the practical courses of general and advanced histopathology had access to both glass-mounted and digitalized slides. Prior to exams, students were asked to answer an anonymous questionnaire. The results of the study reflect the high acceptance and intensive use of the web-based digital histology by students, thus encouraging the development of further Web-based learning strategies for the teaching of histology and pathology. 2010 Elsevier GmbH. All rights reserved.

Phenotype classification of single cells using SRS microscopy, RNA sequencing, and microfluidics (Conference Presentation)

NASA Astrophysics Data System (ADS)

Streets, Aaron M.; Cao, Chen; Zhang, Xiannian; Huang, Yanyi

2016-03-01

Phenotype classification of single cells reveals biological variation that is masked in ensemble measurement. This heterogeneity is found in gene and protein expression as well as in cell morphology. Many techniques are available to probe phenotypic heterogeneity at the single cell level, for example quantitative imaging and single-cell RNA sequencing, but it is difficult to perform multiple assays on the same single cell. In order to directly track correlation between morphology and gene expression at the single cell level, we developed a microfluidic platform for quantitative coherent Raman imaging and immediate RNA sequencing (RNA-Seq) of single cells. With this device we actively sort and trap cells for analysis with stimulated Raman scattering microscopy (SRS). The cells are then processed in parallel pipelines for lysis, and preparation of cDNA for high-throughput transcriptome sequencing. SRS microscopy offers three-dimensional imaging with chemical specificity for quantitative analysis of protein and lipid distribution in single cells. Meanwhile, the microfluidic platform facilitates single-cell manipulation, minimizes contamination, and furthermore, provides improved RNA-Seq detection sensitivity and measurement precision, which is necessary for differentiating biological variability from technical noise. By combining coherent Raman microscopy with RNA sequencing, we can better understand the relationship between cellular morphology and gene expression at the single-cell level.

Use of Raman microscopy and multivariate data analysis to observe the biomimetic growth of carbonated hydroxyapatite on bioactive glass.

PubMed

Seah, Regina K H; Garland, Marc; Loo, Joachim S C; Widjaja, Effendi

2009-02-15

In the present contribution, the biomimetic growth of carbonated hydroxyapatite (HA) on bioactive glass were investigated by Raman microscopy. Bioactive glass samples were immersed in simulated body fluid (SBF) buffered solution at pH 7.40 up to 17 days at 37 degrees C. Raman microscopy mapping was performed on the bioglass samples immersed in SBF solution for different periods of time. The collected data was then analyzed using the band-target entropy minimization technique to extract the observable pure component Raman spectral information. In this study, the pure component Raman spectra of the precursor amorphous calcium phosphate, transient octacalcium phosphate, and matured HA were all recovered. In addition, pure component Raman spectra of calcite, silica glass, and some organic impurities were also recovered. The resolved pure component spectra were fit to the normalized measured Raman data to provide the spatial distribution of these species on the sample surfaces. The current results show that Raman microscopy and multivariate data analysis provide a sensitive and accurate tool to characterize the surface morphology, as well as to give more specific information on the chemical species present and the phase transformation of phosphate species during the formation of HA on bioactive glass.

Challenges of microtome‐based serial block‐face scanning electron microscopy in neuroscience

PubMed Central

WANNER, A. A.; KIRSCHMANN, M. A.

2015-01-01

Summary Serial block‐face scanning electron microscopy (SBEM) is becoming increasingly popular for a wide range of applications in many disciplines from biology to material sciences. This review focuses on applications for circuit reconstruction in neuroscience, which is one of the major driving forces advancing SBEM. Neuronal circuit reconstruction poses exceptional challenges to volume EM in terms of resolution, field of view, acquisition time and sample preparation. Mapping the connections between neurons in the brain is crucial for understanding information flow and information processing in the brain. However, information on the connectivity between hundreds or even thousands of neurons densely packed in neuronal microcircuits is still largely missing. Volume EM techniques such as serial section TEM, automated tape‐collecting ultramicrotome, focused ion‐beam scanning electron microscopy and SBEM (microtome serial block‐face scanning electron microscopy) are the techniques that provide sufficient resolution to resolve ultrastructural details such as synapses and provides sufficient field of view for dense reconstruction of neuronal circuits. While volume EM techniques are advancing, they are generating large data sets on the terabyte scale that require new image processing workflows and analysis tools. In this review, we present the recent advances in SBEM for circuit reconstruction in neuroscience and an overview of existing image processing and analysis pipelines. PMID:25907464

How Hedstrom files fail during clinical use? A retrieval study based on SEM, optical microscopy and micro-XCT analysis.

PubMed

Zinelis, Spiros; Al Jabbari, Youssef S

2018-05-01

This study was conducted to evaluate the failure mechanism of clinically failed Hedstrom (H)-files. Discarded H-files (n=160) from #8 to #40 ISO sizes were collected from different dental clinics. Retrieved files were classified according to their macroscopic appearance and they were investigated under scanning electron microscopy (SEM) and X-ray micro-computed tomography (mXCT). Then the files were embedded in resin along their longitudinal axis and after metallographic grinding and polishing, studied under an incident light microscope. The macroscopic evaluation showed that small ISO sizes (#08-#15) failed by extensive plastic deformation, while larger sizes (≥#20) tended to fracture. Light microscopy and mXCT results coincided showing that unused and plastically deformed files were free of internal defects, while fractured files demonstrate the presence of intense cracking in the flute region. SEM analysis revealed the presence of striations attributed to the fatigue mechanism. Secondary cracks were also identified by optical microscopy and their distribution was correlated to fatigue under bending loading. Experimental results demonstrated that while overloading of cutting instruments is the predominating failure mechanism of small file sizes (#08-#15), fatigue should be considered the fracture mechanism for larger sizes (≥#20).

Quantitative analysis of the effect of environmental-scanning electron microscopy on collagenous tissues.

PubMed

Lee, Woowon; Toussaint, Kimani C

2018-05-31

Environmental-scanning electron microscopy (ESEM) is routinely applied to various biological samples due to its ability to maintain a wet environment while imaging; moreover, the technique obviates the need for sample coating. However, there is limited research carried out on electron-beam (e-beam) induced tissue damage resulting from using the ESEM. In this paper, we use quantitative second-harmonic generation (SHG) microscopy to examine the effects of e-beam exposure from the ESEM on collagenous tissue samples prepared as either fixed, frozen, wet or dehydrated. Quantitative SHG analysis of tissues, before and after ESEM e-beam exposure in low-vacuum mode, reveals evidence of cross-linking of collagen fibers, however there are no structural differences observed in fixed tissue. Meanwhile wet-mode ESEM appears to radically alter the structure from a regular fibrous arrangement to a more random fiber orientation. We also confirm that ESEM images of collagenous tissues show higher spatial resolution compared to SHG microscopy, but the relative tradeoff with collagen specificity reduces its effectiveness in quantifying collagen fiber organization. Our work provides insight on both the limitations of the ESEM for tissue imaging, and the potential opportunity to use as a complementary technique when imaging fine features in the non-collagenous regions of tissue samples.

Segmentation-based retrospective shading correction in fluorescence microscopy E. coli images for quantitative analysis

NASA Astrophysics Data System (ADS)

Mai, Fei; Chang, Chunqi; Liu, Wenqing; Xu, Weichao; Hung, Yeung S.

2009-10-01

Due to the inherent imperfections in the imaging process, fluorescence microscopy images often suffer from spurious intensity variations, which is usually referred to as intensity inhomogeneity, intensity non uniformity, shading or bias field. In this paper, a retrospective shading correction method for fluorescence microscopy Escherichia coli (E. Coli) images is proposed based on segmentation result. Segmentation and shading correction are coupled together, so we iteratively correct the shading effects based on segmentation result and refine the segmentation by segmenting the image after shading correction. A fluorescence microscopy E. Coli image can be segmented (based on its intensity value) into two classes: the background and the cells, where the intensity variation within each class is close to zero if there is no shading. Therefore, we make use of this characteristics to correct the shading in each iteration. Shading is mathematically modeled as a multiplicative component and an additive noise component. The additive component is removed by a denoising process, and the multiplicative component is estimated using a fast algorithm to minimize the intra-class intensity variation. We tested our method on synthetic images and real fluorescence E.coli images. It works well not only for visual inspection, but also for numerical evaluation. Our proposed method should be useful for further quantitative analysis especially for protein expression value comparison.

Light microscopy morphological characteristics of the sperm flagellum may be related to axonemal abnormalities.

PubMed

Mitchell, V; Sigala, J; Ballot, C; Jumeau, F; Barbotin, A L; Duhamel, A; Rives, N; Rigot, J M; Escalier, D; Peers, M C

2015-03-01

Although electron microscopy provides a detailed analysis of ultrastructural abnormalities, this technique is not available in all laboratories. We sought to determine whether certain characteristics of the flagellum as assessed by light microscopy were related to axonemal abnormalities. Forty-one patients with an absence of outer dynein arms (type I), a lack of a central complex (type III) and an absence of peripheral doublets (type IV) were studied. Sperm morphology was scored according to David's modified classification. Flagella with an irregular thickness were classified as being of normal length, short or broken. There were correlations between missing outer dynein arms and abnormal, short or coiled flagellum. Type III patients showed the highest flagellar defects (a short (P = 0.0027) or an absent flagellum (P = 0.011)). Just over 68% of the irregular flagella were short in Type III patients, whereas this value was only 34.5% in type I and 26.4% in type IV (P = 0.002). There was a negative correlation between misassembly and spermatozoa of irregular flagella (r = -0.79; P = 0.019). It is concluded that light microscopy analysis of flagellum abnormalities may help provide a correct diagnosis, identify sperm abnormalities with fertility potentials and outcomes in assisted reproduction technologies and assess the genetic risk. © 2014 Blackwell Verlag GmbH.

Second-harmonic patterned polarization-analyzed reflection confocal microscope

NASA Astrophysics Data System (ADS)

Okoro, Chukwuemeka; Toussaint, Kimani C.

2017-08-01

We introduce the second-harmonic patterned polarization-analyzed reflection confocal (SPPARC) microscope-a multimodal imaging platform that integrates Mueller matrix polarimetry with reflection confocal and second-harmonic generation (SHG) microscopy. SPPARC microscopy provides label-free three-dimensional (3-D), SHG-patterned confocal images that lend themselves to spatially dependent, linear polarimetric analysis for extraction of rich polarization information based on the Mueller calculus. To demonstrate its capabilities, we use SPPARC microscopy to analyze both porcine tendon and ligament samples and find differences in both circular degree-of-polarization and depolarization parameters. Moreover, using the collagen-generated SHG signal as an endogenous counterstain, we show that the technique can be used to provide 3-D polarimetric information of the surrounding extrafibrillar matrix plus cells or EFMC region. The unique characteristics of SPPARC microscopy holds strong potential for it to more accurately and quantitatively describe microstructural changes in collagen-rich samples in three spatial dimensions.

Correlative Fluorescence and Electron Microscopy

PubMed Central

Schirra, Randall T.; Zhang, Peijun

2014-01-01

Correlative fluorescence and electron microscopy (CFEM) is a multimodal technique that combines dynamic and localization information from fluorescence methods with ultrastructural data from electron microscopy, to give new information about how cellular components change relative to the spatiotemporal dynamics within their environment. In this review, we will discuss some of the basic techniques and tools of the trade for utilizing this attractive research method, which is becoming a very powerful tool for biology labs. The information obtained from correlative methods has proven to be invaluable in creating consensus between the two types of microscopy, extending the capability of each, and cutting the time and expense associate with using each method separately for comparative analysis. The realization of the advantages of these methods in cell biology have led to rapid improvement in the protocols and have ushered in a new generation of instruments to reach the next level of correlation – integration. PMID:25271959

Understanding the optics to aid microscopy image segmentation.

PubMed

Yin, Zhaozheng; Li, Kang; Kanade, Takeo; Chen, Mei

2010-01-01

Image segmentation is essential for many automated microscopy image analysis systems. Rather than treating microscopy images as general natural images and rushing into the image processing warehouse for solutions, we propose to study a microscope's optical properties to model its image formation process first using phase contrast microscopy as an exemplar. It turns out that the phase contrast imaging system can be relatively well explained by a linear imaging model. Using this model, we formulate a quadratic optimization function with sparseness and smoothness regularizations to restore the "authentic" phase contrast images that directly correspond to specimen's optical path length without phase contrast artifacts such as halo and shade-off. With artifacts removed, high quality segmentation can be achieved by simply thresholding the restored images. The imaging model and restoration method are quantitatively evaluated on two sequences with thousands of cells captured over several days.

A direct electron detector for time-resolved MeV electron microscopy

DOE PAGES

Vecchione, T.; Denes, P.; Jobe, R. K.; ...

2017-03-15

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here in this paper, we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μm spatial resolution and less than 20 analogue-to-digital converter count RMS pixelmore » noise. The unique capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

Microscopy based studies on the interaction of bio-based silver nanoparticles with Bombyx mori Nuclear Polyhedrosis virus.

PubMed

Tamilselvan, Selvaraj; Ashokkumar, Thirunavukkarasu; Govindaraju, Kasivelu

2017-04-01

In the present investigation, silver nanoparticles (AgNPs) interactions with Bombyx mori Nuclear Polyhedrosis virus (BmNPV) were characterized using High-Resolution Scanning Electron Microscopy (HR-SEM), Energy Dispersive X-ray Analysis (EDAX), Transmission Electron Microscopy (TEM), Atomic Force Microcopy (AFM) and Confocal Microscope (CM). HR-SEM study reveals that the biosynthesized AgNPs have interacted with BmNPV and were found on the surface. TEM micrographs of normal and viral polyhedra treated with AgNPs showed that the nanoparticles were accumulated in the membrane and it was noted that some of the AgNPs successfully penetrated the membrane by reaching the capsid of BmNPV. AFM and confocal microscopy studies reveal that the disruption in the shell membrane tends to lose its stability due to exposure of AgNPs to BmNPV. Copyright © 2017 Elsevier B.V. All rights reserved.

Robust Nucleus/Cell Detection and Segmentation in Digital Pathology and Microscopy Images: A Comprehensive Review.

PubMed

Xing, Fuyong; Yang, Lin

2016-01-01

Digital pathology and microscopy image analysis is widely used for comprehensive studies of cell morphology or tissue structure. Manual assessment is labor intensive and prone to interobserver variations. Computer-aided methods, which can significantly improve the objectivity and reproducibility, have attracted a great deal of interest in recent literature. Among the pipeline of building a computer-aided diagnosis system, nucleus or cell detection and segmentation play a very important role to describe the molecular morphological information. In the past few decades, many efforts have been devoted to automated nucleus/cell detection and segmentation. In this review, we provide a comprehensive summary of the recent state-of-the-art nucleus/cell segmentation approaches on different types of microscopy images including bright-field, phase-contrast, differential interference contrast, fluorescence, and electron microscopies. In addition, we discuss the challenges for the current methods and the potential future work of nucleus/cell detection and segmentation.

Chapter 14: Electron Microscopy on Thin Films for Solar Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Romero, Manuel; Abou-Ras, Daniel; Nichterwitz, Melanie

2016-07-22

This chapter overviews the various techniques applied in scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and highlights their possibilities and also limitations. It gives the various imaging and analysis techniques applied on a scanning electron microscope. The chapter shows that imaging is divided into that making use of secondary electrons (SEs) and of backscattered electrons (BSEs), resulting in different contrasts in the images and thus providing information on compositions, microstructures, and surface potentials. Whenever aiming for imaging and analyses at scales of down to the angstroms range, TEM and its related techniques are appropriate tools. In many cases,more » also SEM techniques provide the access to various material properties of the individual layers, not requiring specimen preparation as time consuming as TEM techniques. Finally, the chapter dedicates to cross-sectional specimen preparation for electron microscopy. The preparation decides indeed on the quality of imaging and analyses.« less

A direct electron detector for time-resolved MeV electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vecchione, T.; Denes, P.; Jobe, R. K.

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μmμm spatial resolution and less than 20 analogue-to-digital converter count RMS pixel noise. The uniquemore » capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

A direct electron detector for time-resolved MeV electron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vecchione, T.; Denes, P.; Jobe, R. K.

The introduction of direct electron detectors enabled the structural biology revolution of cryogenic electron microscopy. Direct electron detectors are now expected to have a similarly dramatic impact on time-resolved MeV electron microscopy, particularly by enabling both spatial and temporal jitter correction. Here in this paper, we report on the commissioning of a direct electron detector for time-resolved MeV electron microscopy. The direct electron detector demonstrated MeV single electron sensitivity and is capable of recording megapixel images at 180 Hz. The detector has a 15-bit dynamic range, better than 30-μm spatial resolution and less than 20 analogue-to-digital converter count RMS pixelmore » noise. The unique capabilities of the direct electron detector and the data analysis required to take advantage of these capabilities are presented. The technical challenges associated with generating and processing large amounts of data are also discussed.« less

Analysis of Peroxisome Biogenesis in Pollen by Confocal Microscopy and Transmission Electron Microscopy.

PubMed

Jia, Peng-Fei; Li, Hong-Ju; Yang, Wei-Cai

2017-01-01

Peroxisome is an essential single-membrane bound organelle in most eukaryotic cells and functions in diverse cellular processes. De novo formation, division, and turnover of peroxisomes contribute to its biogenesis, morphology, and population regulation. In plants, peroxisome plays multiple roles, including metabolism, development, and stress response. Defective peroxisome biogenesis and development retard plant growth, adaption, and reproduction. Through tracing the subcellular localization of fluorescent reporter tagged matrix protein of peroxisome, fluorescence microscopy is a reliable and fast way to detect peroxisome biogenesis. Further fine-structural observation of peroxisome by TEM enables researchers to observe the detailed ultrastructure of its morphology and spatial contact with other organelles. Pollen grain is a specialized structure where two small sperm cells are enclosed in the cytoplasm of a large vegetative cell. Two features make pollen grain a good system to study peroxisome biogenesis: indispensable requirement of peroxisome for germination on the stigma and homogeneity. Here, we describe the methods of studying peroxisome biogenesis in Arabidopsis pollen grains by fluorescent live-imaging with confocal laser scanning microscopy (CLSM) and by DAB-staining based transmission electron microscopy (TEM).

Analysis of liquid suspensions using scanning electron microscopy in transmission: estimation of the water film thickness using Monte-Carlo simulations.

PubMed

Xiao, J; Foray, G; Masenelli-Varlot, K

2018-02-01

Environmental scanning electron microscopy (ESEM) allows the observation of liquids under specific conditions of pressure and temperature. Moreover, when working in the transmission mode, that is in scanning transmission electron microscopy (STEM), nano-objects can be analysed inside a liquid. The contrast in the images is mass-thickness dependent as in STEM-in-TEM (transmission electron microscopy) using closed cells. However, in STEM-in-ESEM, as the liquid-vapour equilibrium is kept dynamically, the thickness of the water droplet remains unknown. In this paper, the contrasts measured in the experimental images are compared with calculations using Monte-Carlo simulations in order to estimate the thickness of water. Two examples are given. On gold nanoparticles, the thickness of a thick film can be estimated thanks to a contrast inversion. On core-shell latex particles, the grey level of the shell compared with those of the core and of the water film gives a relatively precise measurement of the water film thickness. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

[application of the analytical transmission electron microscopy techniques for detection, identification and visualization of localization of nanoparticles of titanium and cerium oxides in mammalian cells].

PubMed

Shebanova, A S; Bogdanov, A G; Ismagulova, T T; Feofanov, A V; Semenyuk, P I; Muronets, V I; Erokhina, M V; Onishchenko, G E; Kirpichnikov, M P; Shaitan, K V

2014-01-01

This work represents the results of the study on applicability of the modern methods of analytical transmission electron microscopy for detection, identification and visualization of localization of nanoparticles of titanium and cerium oxides in A549 cell, human lung adenocarcinoma cell line. A comparative analysis of images of the nanoparticles in the cells obtained in the bright field mode of transmission electron microscopy, under dark-field scanning transmission electron microscopy and high-angle annular dark field scanning transmission electron was performed. For identification of nanoparticles in the cells the analytical techniques, energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy, were compared when used in the mode of obtaining energy spectrum from different particles and element mapping. It was shown that the method for electron tomography is applicable to confirm that nanoparticles are localized in the sample but not coated by contamination. The possibilities and fields of utilizing different techniques for analytical transmission electron microscopy for detection, visualization and identification of nanoparticles in the biological samples are discussed.

A Parallel Distributed-Memory Particle Method Enables Acquisition-Rate Segmentation of Large Fluorescence Microscopy Images

PubMed Central

Afshar, Yaser; Sbalzarini, Ivo F.

2016-01-01

Modern fluorescence microscopy modalities, such as light-sheet microscopy, are capable of acquiring large three-dimensional images at high data rate. This creates a bottleneck in computational processing and analysis of the acquired images, as the rate of acquisition outpaces the speed of processing. Moreover, images can be so large that they do not fit the main memory of a single computer. We address both issues by developing a distributed parallel algorithm for segmentation of large fluorescence microscopy images. The method is based on the versatile Discrete Region Competition algorithm, which has previously proven useful in microscopy image segmentation. The present distributed implementation decomposes the input image into smaller sub-images that are distributed across multiple computers. Using network communication, the computers orchestrate the collectively solving of the global segmentation problem. This not only enables segmentation of large images (we test images of up to 1010 pixels), but also accelerates segmentation to match the time scale of image acquisition. Such acquisition-rate image segmentation is a prerequisite for the smart microscopes of the future and enables online data compression and interactive experiments. PMID:27046144

DNA origami-based standards for quantitative fluorescence microscopy.

PubMed

Schmied, Jürgen J; Raab, Mario; Forthmann, Carsten; Pibiri, Enrico; Wünsch, Bettina; Dammeyer, Thorben; Tinnefeld, Philip

2014-01-01

Validating and testing a fluorescence microscope or a microscopy method requires defined samples that can be used as standards. DNA origami is a new tool that provides a framework to place defined numbers of small molecules such as fluorescent dyes or proteins in a programmed geometry with nanometer precision. The flexibility and versatility in the design of DNA origami microscopy standards makes them ideally suited for the broad variety of emerging super-resolution microscopy methods. As DNA origami structures are durable and portable, they can become a universally available specimen to check the everyday functionality of a microscope. The standards are immobilized on a glass slide, and they can be imaged without further preparation and can be stored for up to 6 months. We describe a detailed protocol for the design, production and use of DNA origami microscopy standards, and we introduce a DNA origami rectangle, bundles and a nanopillar as fluorescent nanoscopic rulers. The protocol provides procedures for the design and realization of fluorescent marks on DNA origami structures, their production and purification, quality control, handling, immobilization, measurement and data analysis. The procedure can be completed in 1-2 d.

Quantifying the assembly of multicomponent molecular machines by single-molecule total internal reflection fluorescence microscopy

PubMed Central

Boehm, Elizabeth M.; Subramanyam, Shyamal; Ghoneim, Mohamed; Washington, M. Todd; Spies, Maria

2016-01-01

Large, dynamic macromolecular complexes play essential roles in many cellular processes. Knowing how the components of these complexes associate with one another and undergo structural rearrangements is critical to understanding how they function. Single-molecule total internal reflection fluorescence (TIRF) microscopy is a powerful approach for addressing these fundamental issues. In this article, we first discuss single-molecule TIRF microscopes and strategies to immobilize and fluorescently label macromolecules. We then review the use of single-molecule TIRF microscopy to study the formation of binary macromolecular complexes using one-color imaging and inhibitors. We conclude with a discussion of the use of TIRF microscopy to examine the formation of higher-order (i.e., ternary, quaternary, etc.) complexes using multi-color setups. The focus throughout this article is on experimental design, controls, data acquisition, and data analysis. We hope that single-molecule TIRF microscopy, which has largely been the province of specialists, will soon become as common in the tool box of biophysicists and biochemists as structural approaches has become today. PMID:27793278

A Parallel Distributed-Memory Particle Method Enables Acquisition-Rate Segmentation of Large Fluorescence Microscopy Images.

PubMed

Afshar, Yaser; Sbalzarini, Ivo F

2016-01-01

Modern fluorescence microscopy modalities, such as light-sheet microscopy, are capable of acquiring large three-dimensional images at high data rate. This creates a bottleneck in computational processing and analysis of the acquired images, as the rate of acquisition outpaces the speed of processing. Moreover, images can be so large that they do not fit the main memory of a single computer. We address both issues by developing a distributed parallel algorithm for segmentation of large fluorescence microscopy images. The method is based on the versatile Discrete Region Competition algorithm, which has previously proven useful in microscopy image segmentation. The present distributed implementation decomposes the input image into smaller sub-images that are distributed across multiple computers. Using network communication, the computers orchestrate the collectively solving of the global segmentation problem. This not only enables segmentation of large images (we test images of up to 10(10) pixels), but also accelerates segmentation to match the time scale of image acquisition. Such acquisition-rate image segmentation is a prerequisite for the smart microscopes of the future and enables online data compression and interactive experiments.

Live CLEM imaging to analyze nuclear structures at high resolution.

PubMed

Haraguchi, Tokuko; Osakada, Hiroko; Koujin, Takako

2015-01-01

Fluorescence microscopy (FM) and electron microscopy (EM) are powerful tools for observing molecular components in cells. FM can provide temporal information about cellular proteins and structures in living cells. EM provides nanometer resolution images of cellular structures in fixed cells. We have combined FM and EM to develop a new method of correlative light and electron microscopy (CLEM), called "Live CLEM." In this method, the dynamic behavior of specific molecules of interest is first observed in living cells using fluorescence microscopy (FM) and then cellular structures in the same cell are observed using electron microscopy (EM). Following image acquisition, FM and EM images are compared to enable the fluorescent images to be correlated with the high-resolution images of cellular structures obtained using EM. As this method enables analysis of dynamic events involving specific molecules of interest in the context of specific cellular structures at high resolution, it is useful for the study of nuclear structures including nuclear bodies. Here we describe Live CLEM that can be applied to the study of nuclear structures in mammalian cells.

3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy

PubMed Central

Lerner, Thomas R.; Burden, Jemima J.; Nkwe, David O.; Pelchen-Matthews, Annegret; Domart, Marie-Charlotte; Durgan, Joanne; Weston, Anne; Jones, Martin L.; Peddie, Christopher J.; Carzaniga, Raffaella; Florey, Oliver; Marsh, Mark; Gutierrez, Maximiliano G.

2017-01-01

ABSTRACT The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here, we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy, and 3D image processing and analysis. We demonstrate successful application of the workflow to three studies, each aiming to better understand complex and dynamic biological processes, including bacterial and viral infections of cultured cells and formation of entotic cell-in-cell structures commonly observed in tumours. Our workflow revealed new insight into the replicative niche of Mycobacterium tuberculosis in primary human lymphatic endothelial cells, HIV-1 in human monocyte-derived macrophages, and the composition of the entotic vacuole. The broad application of this 3D CLEM technique will make it a useful addition to the correlative imaging toolbox for biomedical research. PMID:27445312

Applying fluorescence microscopy to the investigation of the behavior of foodborne pathogens on produce

NASA Astrophysics Data System (ADS)

Brandl, Maria T.

2009-05-01

In the past decade, the development of new tools to better visualize microbes at the cellular scale has spurred a renaissance in the application of microscopy to the study of bacteria in their natural environment. This renewed interest in microscopy may be largely attributable to the advent of the confocal laser scanning microscope (CLSM) and to the discovery of the green fluorescent protein. This article provides information about the use of fluorescence microscopy combined with fluorescent labels such as GFP, DsRed, and DNA stains, with immunofluorescence, and with digital image analysis, to examine the behavior of bacteria and other microbes on plant surfaces. Some of the advantages and pitfalls of these methods will be described using practical examples derived from studies of the ecology of foodborne pathogens, namely Salmonella enterica and E. coli O157:H7, on fresh fruit and vegetables. Confocal microscopy has been a powerful approach to uncover some of the factors involved in the association of produce with epidemics caused by these human pathogens and their interaction with other microbes in their nonhost environment.

Filling the gap: adding super-resolution to array tomography for correlated ultrastructural and molecular identification of electrical synapses at the C. elegans connectome.

PubMed

Markert, Sebastian Matthias; Britz, Sebastian; Proppert, Sven; Lang, Marietta; Witvliet, Daniel; Mulcahy, Ben; Sauer, Markus; Zhen, Mei; Bessereau, Jean-Louis; Stigloher, Christian

2016-10-01

Correlating molecular labeling at the ultrastructural level with high confidence remains challenging. Array tomography (AT) allows for a combination of fluorescence and electron microscopy (EM) to visualize subcellular protein localization on serial EM sections. Here, we describe an application for AT that combines near-native tissue preservation via high-pressure freezing and freeze substitution with super-resolution light microscopy and high-resolution scanning electron microscopy (SEM) analysis on the same section. We established protocols that combine SEM with structured illumination microscopy (SIM) and direct stochastic optical reconstruction microscopy (dSTORM). We devised a method for easy, precise, and unbiased correlation of EM images and super-resolution imaging data using endogenous cellular landmarks and freely available image processing software. We demonstrate that these methods allow us to identify and label gap junctions in Caenorhabditis elegans with precision and confidence, and imaging of even smaller structures is feasible. With the emergence of connectomics, these methods will allow us to fill in the gap-acquiring the correlated ultrastructural and molecular identity of electrical synapses.

Are rapid diagnostic tests more accurate in diagnosis of plasmodium falciparum malaria compared to microscopy at rural health centres?

PubMed

Batwala, Vincent; Magnussen, Pascal; Nuwaha, Fred

2010-12-02

Prompt, accurate diagnosis and treatment with artemisinin combination therapy remains vital to current malaria control. Blood film microscopy the current standard test for diagnosis of malaria has several limitations that necessitate field evaluation of alternative diagnostic methods especially in low income countries of sub-Saharan Africa where malaria is endemic. The accuracy of axillary temperature, health centre (HC) microscopy, expert microscopy and a HRP2-based rapid diagnostic test (Paracheck) was compared in predicting malaria infection using polymerase chain reaction (PCR) as the gold standard. Three hundred patients with a clinical suspicion of malaria based on fever and or history of fever from a low and high transmission setting in Uganda were consecutively enrolled and provided blood samples for all tests. Accuracy of each test was calculated overall with 95% confidence interval and then adjusted for age-groups and level of transmission intensity using a stratified analysis. The endpoints were: sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV). This study is registered with Clinicaltrials.gov, NCT00565071. Of the 300 patients, 88(29.3%) had fever, 56(18.7%) were positive by HC microscopy, 47(15.7%) by expert microscopy, 110(36.7%) by Paracheck and 89(29.7%) by PCR. The overall sensitivity >90% was only shown by Paracheck 91.0% [95%CI: 83.1-96.0]. The sensitivity of expert microscopy was 46%, similar to HC microscopy. The superior sensitivity of Paracheck compared to microscopy was maintained when data was stratified for transmission intensity and age. The overall specificity rates were: Paracheck 86.3% [95%CI: 80.9-90.6], HC microscopy 93.4% [95%CI: 89.1-96.3] and expert microscopy 97.2% [95%CI: 93.9-98.9]. The NPV >90% was shown by Paracheck 95.8% [95%CI: 91.9-98.2]. The overall PPV was <88% for all methods. The HRP2-based RDT has shown superior sensitivity compared to microscopy in diagnosis of malaria and may be more suitable for screening of malaria infection.

Chemical imaging of molecular changes in a hydrated single cell by dynamic secondary ion mass spectrometry and super-resolution microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hua, Xin; Szymanski, Craig; Wang, Zhaoying

2016-01-01

Chemical imaging of single cells is important in capturing biological dynamics. Single cell correlative imaging is realized between structured illumination microscopy (SIM) and time-of-flight secondary ion mass spectrometry (ToF-SIMS) using System for Analysis at the Liquid Vacuum Interface (SALVI), a multimodal microreactor. SIM characterized cells and guided subsequent ToF-SIMS analysis. Dynamic ToF-SIMS provided time- and space-resolved cell molecular mapping. Lipid fragments were identified in the hydrated cell membrane. Principal component analysis was used to elucidate chemical component differences among mouse lung cells that uptake zinc oxide nanoparticles. Our results provided submicron chemical spatial mapping for investigations of cell dynamics atmore » the molecular level.« less

Probing of multiple magnetic responses in magnetic inductors using atomic force microscopy.

PubMed

Park, Seongjae; Seo, Hosung; Seol, Daehee; Yoon, Young-Hwan; Kim, Mi Yang; Kim, Yunseok

2016-02-08

Even though nanoscale analysis of magnetic properties is of significant interest, probing methods are relatively less developed compared to the significance of the technique, which has multiple potential applications. Here, we demonstrate an approach for probing various magnetic properties associated with eddy current, coil current and magnetic domains in magnetic inductors using multidimensional magnetic force microscopy (MMFM). The MMFM images provide combined magnetic responses from the three different origins, however, each contribution to the MMFM response can be differentiated through analysis based on the bias dependence of the response. In particular, the bias dependent MMFM images show locally different eddy current behavior with values dependent on the type of materials that comprise the MI. This approach for probing magnetic responses can be further extended to the analysis of local physical features.

Analysing magnetism using scanning SQUID microscopy.

PubMed

Reith, P; Renshaw Wang, X; Hilgenkamp, H

2017-12-01

Scanning superconducting quantum interference device microscopy (SSM) is a scanning probe technique that images local magnetic flux, which allows for mapping of magnetic fields with high field and spatial accuracy. Many studies involving SSM have been published in the last few decades, using SSM to make qualitative statements about magnetism. However, quantitative analysis using SSM has received less attention. In this work, we discuss several aspects of interpreting SSM images and methods to improve quantitative analysis. First, we analyse the spatial resolution and how it depends on several factors. Second, we discuss the analysis of SSM scans and the information obtained from the SSM data. Using simulations, we show how signals evolve as a function of changing scan height, SQUID loop size, magnetization strength, and orientation. We also investigated 2-dimensional autocorrelation analysis to extract information about the size, shape, and symmetry of magnetic features. Finally, we provide an outlook on possible future applications and improvements.

Analysing magnetism using scanning SQUID microscopy

NASA Astrophysics Data System (ADS)

Reith, P.; Renshaw Wang, X.; Hilgenkamp, H.

2017-12-01

Scanning superconducting quantum interference device microscopy (SSM) is a scanning probe technique that images local magnetic flux, which allows for mapping of magnetic fields with high field and spatial accuracy. Many studies involving SSM have been published in the last few decades, using SSM to make qualitative statements about magnetism. However, quantitative analysis using SSM has received less attention. In this work, we discuss several aspects of interpreting SSM images and methods to improve quantitative analysis. First, we analyse the spatial resolution and how it depends on several factors. Second, we discuss the analysis of SSM scans and the information obtained from the SSM data. Using simulations, we show how signals evolve as a function of changing scan height, SQUID loop size, magnetization strength, and orientation. We also investigated 2-dimensional autocorrelation analysis to extract information about the size, shape, and symmetry of magnetic features. Finally, we provide an outlook on possible future applications and improvements.

Electrical relaxation, optical and magnetic studies of nanocrystalline lithium ferrite synthesized by different chemical routes

NASA Astrophysics Data System (ADS)

Cheruku, Rajesh; Govindaraj, G.; Vijayan, Lakshmi

2017-12-01

The nanocrystalline lithium ferrite was synthesized by wet chemical methods such as solution combustion technique, sol-gel, and hydrothermal for a comparative study. Different characterization techniques like x-ray powder diffraction and thermal analysis were employed to confirm the structure and phase. Temperature-dependent Raman analysis was employed to classify the phonon modes associated with precise atomic motions existing in the synthesized materials. Morphology of sample surface was explored by scanning electron microscopy, and elemental analysis was done by energy dispersive spectroscopy analysis. The nanocrystalline nature of the materials was confirmed through transmission electron microscopy. Magnetic properties of these samples were explored through a vibrating sample magnetometer. Ac electrical impedance spectroscopy data were investigated using two Cole-Cole functions, and activation energies were calculated for all materials. Among them, solution combustion prepared lithium ferrite shows the highest conductivity and lowest activation energy.

Biosynthesis of silver nanoparticles by a Bacillus sp. of marine origin

NASA Astrophysics Data System (ADS)

Janardhanan, A.; Roshmi, T.; Varghese, Rintu T.; Soniya, E. V.; Mathew, Jyothis; Radhakrishnan, E. K.

2013-04-01

This study was aimed to explore the nanoparticle synthesizing properties of a silver resistant Bacillus sp. isolated from a marine water sample. The 16SrDNA sequence analysis of the isolate proved it as a Bacillus strain. Very interestingly, the isolate was found to have the ability to form intracellular silver nanoparticles at room temperature within 24 hours. This was confirmed by the UV-Vis absorption analysis which showed a peak at 430 nm corresponding to the plasmon absorbance of silver nanoparticles. Further characterization of the nanoparticles was carried out by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) analysis. The presence of silver nanoparticles with the size less than 100 nm was confirmed. These particles were found to be extremely stable as confirmed by the TEM analysis after three months of purification. So, the current study is the demonstration of an efficient synthesis of stable silver nanoparticles by a marine Bacillus strain.

Comparative analysis of the modified enclosed energy metric for self-focusing holograms from digital lensless holographic microscopy.

PubMed

Trujillo, Carlos; Garcia-Sucerquia, Jorge

2015-06-01

A comparative analysis of the performance of the modified enclosed energy (MEE) method for self-focusing holograms recorded with digital lensless holographic microscopy is presented. Notwithstanding the MEE analysis previously published, no extended analysis of its performance has been reported. We have tested the MEE in terms of the minimum axial distance allowed between the set of reconstructed holograms to search for the focal plane and the elapsed time to obtain the focused image. These parameters have been compared with those for some of the already reported methods in the literature. The MEE achieves better results in terms of self-focusing quality but at a higher computational cost. Despite its longer processing time, the method remains within a time frame to be technologically attractive. Modeled and experimental holograms have been utilized in this work to perform the comparative study.

Quantitative nanoscopy: Tackling sampling limitations in (S)TEM imaging of polymers and composites.

PubMed

Gnanasekaran, Karthikeyan; Snel, Roderick; de With, Gijsbertus; Friedrich, Heiner

2016-01-01

Sampling limitations in electron microscopy questions whether the analysis of a bulk material is representative, especially while analyzing hierarchical morphologies that extend over multiple length scales. We tackled this problem by automatically acquiring a large series of partially overlapping (S)TEM images with sufficient resolution, subsequently stitched together to generate a large-area map using an in-house developed acquisition toolbox (TU/e Acquisition ToolBox) and stitching module (TU/e Stitcher). In addition, we show that quantitative image analysis of the large scale maps provides representative information that can be related to the synthesis and process conditions of hierarchical materials, which moves electron microscopy analysis towards becoming a bulk characterization tool. We demonstrate the power of such an analysis by examining two different multi-phase materials that are structured over multiple length scales. Copyright © 2015 Elsevier B.V. All rights reserved.

Lens-free microscopy of cerebrospinal fluid for the laboratory diagnosis of meningitis

NASA Astrophysics Data System (ADS)

Delacroix, Robin; Morel, Sophie Nhu An; Hervé, Lionel; Bordy, Thomas; Blandin, Pierre; Dinten, Jean-Marc; Drancourt, Michel; Allier, Cédric

2018-02-01

The cytology of the cerebrospinal fluid is traditionally performed by an operator (physician, biologist) by means of a conventional light microscope. The operator visually counts the leukocytes (white blood cells) present in a sample of cerebrospinal fluid (10 μl). It is a tedious job and the result is operator-dependent. Here in order to circumvent the limitations of manual counting, we approach the question of numeration of erythrocytes and leukocytes for the cytological diagnosis of meningitis by means of lens-free microscopy. In a first step, a prospective counts of leukocytes was performed by five different operators using conventional optical microscopy. The visual counting yielded an overall 16.7% misclassification of 72 cerebrospinal fluid specimens in meningitis/non-meningitis categories using a 10 leukocyte/μL cut-off. In a second step, the lens-free microscopy algorithm was adapted step-by-step for counting cerebrospinal fluid cells and discriminating leukocytes from erythrocytes. The optimization of the automatic lens-free counting was based on the prospective analysis of 215 cerebrospinal fluid specimens. The optimized algorithm yielded a 100% sensitivity and a 86% specificity compared to confirmed diagnostics. In a third step, a blind lens-free microscopic analysis of 116 cerebrospinal fluid specimens, including six cases of microbiology confirmed infectious meningitis, yielded a 100% sensitivity and a 79% specificity. Adapted lens-free microscopy is thus emerging as an operator-independent technique for the rapid numeration of leukocytes and erythrocytes in cerebrospinal fluid. In particular, this technique is well suited to the rapid diagnosis of meningitis at point-of-care laboratories.

Label-free three-dimensional imaging of cell nucleus using third-harmonic generation microscopy

NASA Astrophysics Data System (ADS)

Lin, Jian; Zheng, Wei; Wang, Zi; Huang, Zhiwei

2014-09-01

We report the implementation of the combined third-harmonic generation (THG) and two-photon excited fluorescence (TPEF) microscopy for label-free three-dimensional (3-D) imaging of cell nucleus morphological changes in liver tissue. THG imaging shows regular spherical shapes of normal hepatocytes nuclei with inner chromatin structures while revealing the condensation of chromatins and nuclear fragmentations in hepatocytes of diseased liver tissue. Colocalized THG and TPEF imaging provides complementary information of cell nuclei and cytoplasm in tissue. This work suggests that 3-D THG microscopy has the potential for quantitative analysis of nuclear morphology in cells at a submicron-resolution without the need for DNA staining.

Comparison of the morphology, chemical composition and microstructure of cryptocrystalline graphite and carbon black

NASA Astrophysics Data System (ADS)

Quan, Ying; Liu, Qinfu; Zhang, Shilong; Zhang, Shuai

2018-07-01

The structures of cryptocrystalline graphite (CG) and carbon black (CB) have been analyzed using scanning electron microscopy (SEM), transmission electron microscopy (TEM), organic elemental analysis (OEA), X-ray diffraction (XRD), RAMAN and high-resolution transmission electron microscopy (HRTEM). These results indicate that CG has the same elemental composition as CB, with carbon being the major element present. SL sample (CG with low graphitization degree) and CB exhibit similar microcrystalline structures. CG was shown to contain a layered graphitic structure that was significantly different to the primary spherical particles present in CB. It is proposed that these CG sheets may potentially be reduced and delaminated to afford multilayer graphene structures with improved material properties.

Label-free three-dimensional imaging of cell nucleus using third-harmonic generation microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Jian; Zheng, Wei; Wang, Zi

2014-09-08

We report the implementation of the combined third-harmonic generation (THG) and two-photon excited fluorescence (TPEF) microscopy for label-free three-dimensional (3-D) imaging of cell nucleus morphological changes in liver tissue. THG imaging shows regular spherical shapes of normal hepatocytes nuclei with inner chromatin structures while revealing the condensation of chromatins and nuclear fragmentations in hepatocytes of diseased liver tissue. Colocalized THG and TPEF imaging provides complementary information of cell nuclei and cytoplasm in tissue. This work suggests that 3-D THG microscopy has the potential for quantitative analysis of nuclear morphology in cells at a submicron-resolution without the need for DNA staining.

Mineralogy and Microstructures of Shock-Induced Melt Veins in Chondrites

NASA Technical Reports Server (NTRS)

Sharp, Thomas G.

2000-01-01

The applicability of phase equilibrium data to the interpretation of shock-induced melt veins can only be tested by a detailed study of melt- vein mineralogy to see how high-pressure assemblages vary as a function of shock conditions inferred from other indicators. We have used transmission electron microscopy (TEM), analytical electron microscopy (AEM), scanning electron microscopy (SEM), electron microprobe analysis (EMA) and optical petrography to characterize the mineralogy, microstructures, and compositions of melt veins and associated high-pressure minerals in shocked chondrites and SNC meteorites. In the processes, we have gained a better understanding of what melt veining can tell us about shock conditions and we have discovered new mineral phases in chondritic and SNC meteorites.

Even illumination in total internal reflection fluorescence microscopy using laser light.

PubMed

Fiolka, R; Belyaev, Y; Ewers, H; Stemmer, A

2008-01-01

In modern fluorescence microscopy, lasers are a widely used source of light, both for imaging in total internal reflection and epi-illumination modes. In wide-field imaging, scattering of highly coherent laser light due to imperfections in the light path typically leads to nonuniform illumination of the specimen, compromising image analysis. We report the design and construction of an objective-launch total internal reflection fluorescence microscopy system with excellent evenness of specimen illumination achieved by azimuthal rotation of the incoming illuminating laser beam. The system allows quick and precise changes of the incidence angle of the laser beam and thus can also be used in an epifluorescence mode. 2007 Wiley-Liss, Inc

Nanopore fabrication and characterization by helium ion microscopy

NASA Astrophysics Data System (ADS)

Emmrich, D.; Beyer, A.; Nadzeyka, A.; Bauerdick, S.; Meyer, J. C.; Kotakoski, J.; Gölzhäuser, A.

2016-04-01

The Helium Ion Microscope (HIM) has the capability to image small features with a resolution down to 0.35 nm due to its highly focused gas field ionization source and its small beam-sample interaction volume. In this work, the focused helium ion beam of a HIM is utilized to create nanopores with diameters down to 1.3 nm. It will be demonstrated that nanopores can be milled into silicon nitride, carbon nanomembranes, and graphene with well-defined aspect ratio. To image and characterize the produced nanopores, helium ion microscopy and high resolution scanning transmission electron microscopy were used. The analysis of the nanopores' growth behavior allows inferring on the profile of the helium ion beam.

Automated microscopy for high-content RNAi screening

PubMed Central

2010-01-01

Fluorescence microscopy is one of the most powerful tools to investigate complex cellular processes such as cell division, cell motility, or intracellular trafficking. The availability of RNA interference (RNAi) technology and automated microscopy has opened the possibility to perform cellular imaging in functional genomics and other large-scale applications. Although imaging often dramatically increases the content of a screening assay, it poses new challenges to achieve accurate quantitative annotation and therefore needs to be carefully adjusted to the specific needs of individual screening applications. In this review, we discuss principles of assay design, large-scale RNAi, microscope automation, and computational data analysis. We highlight strategies for imaging-based RNAi screening adapted to different library and assay designs. PMID:20176920

Femtosecond laser subsurface scleral treatment in cadaver human sclera and evaluation using two-photon and confocal microscopy

NASA Astrophysics Data System (ADS)

Sun, Hui; Fan, Zhongwei; Yan, Ying; Lian, Fuqiang; Kurtz, Ron; Juhasz, Tibor

2016-03-01

Glaucoma is the second-leading cause of blindness worldwide and is often associated with elevated intraocular pressure (IOP). Partial-thickness drainage channels can be created with femtosecond laser in the translucent sclera for the potential treatment of glaucoma. We demonstrate the creation of partial-thickness subsurface drainage channels with the femtosecond laser in the cadaver human eyeballs and describe the application of two-photon microscopy and confocal microscopy for noninvasive imaging of the femtosecond laser created partial-thickness scleral channels in cadaver human eyes. A femtosecond laser operating at a wavelength of 1700 nm was scanned along a rectangular raster pattern to create the partial thickness subsurface drainage channels in the sclera of cadaver human eyes. Analysis of the dimensions and location of these channels is important in understanding their effects. We describe the application of two-photon microscopy and confocal microscopy for noninvasive imaging of the femtosecond laser created partial-thickness scleral channels in cadaver human eyes. High-resolution images, hundreds of microns deep in the sclera, were obtained to allow determination of the shape and dimension of such partial thickness subsurface scleral channels. Our studies suggest that the confocal and two-photon microscopy can be used to investigate femtosecond-laser created partial-thickness drainage channels in the sclera of cadaver human eyes.

Transmission electron microscopy, fluorescence microscopy, and confocal raman microscopic analysis of ultrastructural and compositional heterogeneity of Cornus alba L. wood cell wall.

PubMed

Ma, Jianfeng; Ji, Zhe; Zhou, Xia; Zhang, Zhiheng; Xu, Feng

2013-02-01

Transmission electron microscopy (TEM), fluorescence microscopy, and confocal Raman microscopy can be used to characterize ultrastructural and compositional heterogeneity of plant cell walls. In this study, TEM observations revealed the ultrastructural characterization of Cornus alba L. fiber, vessel, axial parenchyma, ray parenchyma, and pit membrane between cells, notably with the ray parenchyma consisting of two well-defined layers. Fluorescence microscopy evidenced that cell corner middle lamella was more lignified than adjacent compound middle lamella and secondary wall with variation in lignification level from cell to cell. In situ Raman images showed that the inhomogeneity in cell wall components (cellulose and lignin) among different cells and within morphologically distinct cell wall layers. As the significant precursors of lignin biosynthesis, the pattern of coniferyl alcohol and aldehyde (joint abbreviation Lignin-CAA for both structures) distribution in fiber cell wall was also identified by Raman images, with higher concentration occurring in the fiber secondary wall where there was the highest cellulose concentration. Moreover, noteworthy was the observation that higher concentration of lignin and very minor amounts of cellulose were visualized in the pit membrane areas. These complementary microanalytical methods provide more accurate and complete information with regard to ultrastructural and compositional characterization of plant cell walls.

An overview of the legislation and light microscopy for detection of processed animal proteins in feeds.

PubMed

Liu, Xian; Han, Lujia; Veys, Pascal; Baeten, Vincent; Jiang, Xunpeng; Dardenne, Pierre

2011-08-01

From the first cases of bovine spongiform encephalopathy (BSE) among cattle in the United Kingdom in 1986, the route of infection of BSE is generally believed by means of feeds containing low level of processed animal proteins (PAPs). Therefore, many feed bans and alternative and complementary techniques were resulted for the BSE safeguards in the world. Now the feed bans are expected to develop into a "species to species" ban, which requires the corresponding species-specific identification methods. Currently, banned PAPs can be detected by various methods as light microscopy, polymerase chain reaction, enzyme-linked immunosorbent assay, near infrared spectroscopy, and near infrared microscopy. Light microscopy as described in the recent Commission Regulation EC/152/2009 is the only official method for the detection and characterization of PAPs in feed in the European Union. It is able to detect the presence of constituents of animal origin in feed at the level of 1 g/kg with hardly any false negative. Nevertheless, light microscopy has the limitation of lack of species specificity. This article presents a review of legislations on the use of PAPs in feedstuff, the detection details of animal proteins by light microscopy, and also presents and discusses the analysis procedure and expected development of the technique. Copyright © 2010 Wiley-Liss, Inc.

Limitations of using Raman microscopy for the analysis of high-content-carbon-filled ethylene propylene diene monomer rubber.

PubMed

Ghanbari-Siahkali, Afshin; Almdal, Kristoffer; Kingshott, Peter

2003-12-01

The effects of laser irradiation on changes to the surface chemistry and structure of a commercially available ethylene propylene diene monomer (EPDM) rubber sample after Raman microscopy analysis was investigated. The Raman measurements were carried out with different levels of laser power on the sample, ranging from 4.55 mW to 0.09 mW. The surface of the EPDM was analyzed before and after laser exposure using X-ray photoelectron spectroscopy (XPS) and attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. The techniques have surface probe depths of approximately < or = 10 nm and 1 microm, respectively. Both sets of analysis show that ingredients of the blended EPDM rubber "bloom" to the surface as a result of local heating that takes place due to the absorption of laser by carbon black during the Raman analysis. Scanning electron microscopy (SEM) analysis was also performed on the Raman analyzed areas to visually illustrate the effects created due to laser light exposure (i.e., burning marks). The change in surface chemistry also occurs in regions a few millimeters from the exposed sites, indicating that the effect is quite long range. However, this phenomenon has no major influence, as far as XPS or ATR-FTIR results disclose, on the backbone structure of the rubber sample. The results indicate that precautions should be taken when analyzing complex blended polymer samples using Raman spectroscopy.

Whole Slide Imaging Versus Microscopy for Primary Diagnosis in Surgical Pathology

PubMed Central

Mukhopadhyay, Sanjay; Feldman, Michael D.; Abels, Esther; Ashfaq, Raheela; Beltaifa, Senda; Cacciabeve, Nicolas G.; Cathro, Helen P.; Cheng, Liang; Cooper, Kumarasen; Dickey, Glenn E.; Gill, Ryan M.; Heaton, Robert P.; Kerstens, René; Lindberg, Guy M.; Malhotra, Reenu K.; Mandell, James W.; Manlucu, Ellen D.; Mills, Anne M.; Mills, Stacey E.; Moskaluk, Christopher A.; Nelis, Mischa; Patil, Deepa T.; Przybycin, Christopher G.; Reynolds, Jordan P.; Rubin, Brian P.; Saboorian, Mohammad H.; Salicru, Mauricio; Samols, Mark A.; Sturgis, Charles D.; Turner, Kevin O.; Wick, Mark R.; Yoon, Ji Y.; Zhao, Po

2018-01-01

Most prior studies of primary diagnosis in surgical pathology using whole slide imaging (WSI) versus microscopy have focused on specific organ systems or included relatively few cases. The objective of this study was to demonstrate that WSI is noninferior to microscopy for primary diagnosis in surgical pathology. A blinded randomized noninferiority study was conducted across the entire range of surgical pathology cases (biopsies and resections, including hematoxylin and eosin, immunohistochemistry, and special stains) from 4 institutions using the original sign-out diagnosis (baseline diagnosis) as the reference standard. Cases were scanned, converted to WSI and randomized. Sixteen pathologists interpreted cases by microscopy or WSI, followed by a wash-out period of ≥4 weeks, after which cases were read by the same observers using the other modality. Major discordances were identified by an adjudication panel, and the differences between major discordance rates for both microscopy (against the reference standard) and WSI (against the reference standard) were calculated. A total of 1992 cases were included, resulting in 15,925 reads. The major discordance rate with the reference standard diagnosis was 4.9% for WSI and 4.6% for microscopy. The difference between major discordance rates for microscopy and WSI was 0.4% (95% confidence interval, −0.30% to 1.01%). The difference in major discordance rates for WSI and microscopy was highest in endocrine pathology (1.8%), neoplastic kidney pathology (1.5%), urinary bladder pathology (1.3%), and gynecologic pathology (1.2%). Detailed analysis of these cases revealed no instances where interpretation by WSI was consistently inaccurate compared with microscopy for multiple observers. We conclude that WSI is noninferior to microscopy for primary diagnosis in surgical pathology, including biopsies and resections stained with hematoxylin and eosin, immunohistochemistry and special stains. This conclusion is valid across a wide variety of organ systems and specimen types. PMID:28961557

Whole Slide Imaging Versus Microscopy for Primary Diagnosis in Surgical Pathology: A Multicenter Blinded Randomized Noninferiority Study of 1992 Cases (Pivotal Study).

PubMed

Mukhopadhyay, Sanjay; Feldman, Michael D; Abels, Esther; Ashfaq, Raheela; Beltaifa, Senda; Cacciabeve, Nicolas G; Cathro, Helen P; Cheng, Liang; Cooper, Kumarasen; Dickey, Glenn E; Gill, Ryan M; Heaton, Robert P; Kerstens, René; Lindberg, Guy M; Malhotra, Reenu K; Mandell, James W; Manlucu, Ellen D; Mills, Anne M; Mills, Stacey E; Moskaluk, Christopher A; Nelis, Mischa; Patil, Deepa T; Przybycin, Christopher G; Reynolds, Jordan P; Rubin, Brian P; Saboorian, Mohammad H; Salicru, Mauricio; Samols, Mark A; Sturgis, Charles D; Turner, Kevin O; Wick, Mark R; Yoon, Ji Y; Zhao, Po; Taylor, Clive R

2018-01-01

Most prior studies of primary diagnosis in surgical pathology using whole slide imaging (WSI) versus microscopy have focused on specific organ systems or included relatively few cases. The objective of this study was to demonstrate that WSI is noninferior to microscopy for primary diagnosis in surgical pathology. A blinded randomized noninferiority study was conducted across the entire range of surgical pathology cases (biopsies and resections, including hematoxylin and eosin, immunohistochemistry, and special stains) from 4 institutions using the original sign-out diagnosis (baseline diagnosis) as the reference standard. Cases were scanned, converted to WSI and randomized. Sixteen pathologists interpreted cases by microscopy or WSI, followed by a wash-out period of ≥4 weeks, after which cases were read by the same observers using the other modality. Major discordances were identified by an adjudication panel, and the differences between major discordance rates for both microscopy (against the reference standard) and WSI (against the reference standard) were calculated. A total of 1992 cases were included, resulting in 15,925 reads. The major discordance rate with the reference standard diagnosis was 4.9% for WSI and 4.6% for microscopy. The difference between major discordance rates for microscopy and WSI was 0.4% (95% confidence interval, -0.30% to 1.01%). The difference in major discordance rates for WSI and microscopy was highest in endocrine pathology (1.8%), neoplastic kidney pathology (1.5%), urinary bladder pathology (1.3%), and gynecologic pathology (1.2%). Detailed analysis of these cases revealed no instances where interpretation by WSI was consistently inaccurate compared with microscopy for multiple observers. We conclude that WSI is noninferior to microscopy for primary diagnosis in surgical pathology, including biopsies and resections stained with hematoxylin and eosin, immunohistochemistry and special stains. This conclusion is valid across a wide variety of organ systems and specimen types.

Polarized Light Microscopy

NASA Technical Reports Server (NTRS)

Frandsen, Athela F.

2016-01-01

Polarized light microscopy (PLM) is a technique which employs the use of polarizing filters to obtain substantial optical property information about the material which is being observed. This information can be combined with other microscopy techniques to confirm or elucidate the identity of an unknown material, determine whether a particular contaminant is present (as with asbestos analysis), or to provide important information that can be used to refine a manufacturing or chemical process. PLM was the major microscopy technique in use for identification of materials for nearly a century since its introduction in 1834 by William Fox Talbot, as other techniques such as SEM (Scanning Electron Microscopy), FTIR (Fourier Transform Infrared spectroscopy), XPD (X-ray Powder Diffraction), and TEM (Transmission Electron Microscopy) had not yet been developed. Today, it is still the only technique approved by the Environmental Protection Agency (EPA) for asbestos analysis, and is often the technique first applied for identification of unknown materials. PLM uses different configurations in order to determine different material properties. With each configuration additional clues can be gathered, leading to a conclusion of material identity. With no polarizing filter, the microscope can be used just as a stereo optical microscope, and view qualities such as morphology, size, and number of phases. With a single polarizing filter (single polars), additional properties can be established, such as pleochroism, individual refractive indices, and dispersion staining. With two polarizing filters (crossed polars), even more can be deduced: isotropy vs. anisotropy, extinction angle, birefringence/degree of birefringence, sign of elongation, and anomalous polarization colors, among others. With the use of PLM many of these properties can be determined in a matter of seconds, even for those who are not highly trained. McCrone, a leader in the field of polarized light microscopy, often advised, If you cant determine a specific optical property of a particle after two minutes, move onto another configuration. Since optical properties can be seen so very quickly and easily under polarized light, it is only necessary to spend a maximum of two minutes on a technique to determine a particular property, though often only a few seconds are required.

Analysis of acute brain slices by electron microscopy: a correlative light-electron microscopy workflow based on Tokuyasu cryo-sectioning.

PubMed

Loussert Fonta, Celine; Leis, Andrew; Mathisen, Cliff; Bouvier, David S; Blanchard, Willy; Volterra, Andrea; Lich, Ben; Humbel, Bruno M

2015-01-01

Acute brain slices are slices of brain tissue that are kept vital in vitro for further recordings and analyses. This tool is of major importance in neurobiology and allows the study of brain cells such as microglia, astrocytes, neurons and their inter/intracellular communications via ion channels or transporters. In combination with light/fluorescence microscopies, acute brain slices enable the ex vivo analysis of specific cells or groups of cells inside the slice, e.g. astrocytes. To bridge ex vivo knowledge of a cell with its ultrastructure, we developed a correlative microscopy approach for acute brain slices. The workflow begins with sampling of the tissue and precise trimming of a region of interest, which contains GFP-tagged astrocytes that can be visualised by fluorescence microscopy of ultrathin sections. The astrocytes and their surroundings are then analysed by high resolution scanning transmission electron microscopy (STEM). An important aspect of this workflow is the modification of a commercial cryo-ultramicrotome to observe the fluorescent GFP signal during the trimming process. It ensured that sections contained at least one GFP astrocyte. After cryo-sectioning, a map of the GFP-expressing astrocytes is established and transferred to correlation software installed on a focused ion beam scanning electron microscope equipped with a STEM detector. Next, the areas displaying fluorescence are selected for high resolution STEM imaging. An overview area (e.g. a whole mesh of the grid) is imaged with an automated tiling and stitching process. In the final stitched image, the local organisation of the brain tissue can be surveyed or areas of interest can be magnified to observe fine details, e.g. vesicles or gold labels on specific proteins. The robustness of this workflow is contingent on the quality of sample preparation, based on Tokuyasu's protocol. This method results in a reasonable compromise between preservation of morphology and maintenance of antigenicity. Finally, an important feature of this approach is that the fluorescence of the GFP signal is preserved throughout the entire preparation process until the last step before electron microscopy. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Hierarchical super-structure identified by polarized light microscopy, electron microscopy and nanoindentation: Implications for the limits of biological control over the growth mode of abalone sea shells

PubMed Central

2012-01-01

Background Mollusc shells are commonly investigated using high-resolution imaging techniques based on cryo-fixation. Less detailed information is available regarding the light-optical properties. Sea shells of Haliotis pulcherina were embedded for polishing in defined orientations in order to investigate the interface between prismatic calcite and nacreous aragonite by standard materialographic methods. A polished thin section of the interface was prepared with a defined thickness of 60 μm for quantitative birefringence analysis using polarized light and LC-PolScope microscopy. Scanning electron microscopy images were obtained for comparison. In order to study structural-mechanical relationships, nanoindentation experiments were performed. Results Incident light microscopy revealed a super-structure in semi-transparent regions of the polished cross-section under a defined angle. This super-structure is not visible in transmitted birefringence analysis due to the blurred polarization of small nacre platelets and numerous organic interfaces. The relative orientation and homogeneity of calcite prisms was directly identified, some of them with their optical axes exactly normal to the imaging plane. Co-oriented "prism colonies" were identified by polarized light analyses. The nacreous super-structure was also visualized by secondary electron imaging under defined angles. The domains of the super-structure were interpreted to consist of crystallographically aligned platelet stacks. Nanoindentation experiments showed that mechanical properties changed with the same periodicity as the domain size. Conclusions In this study, we have demonstrated that insights into the growth mechanisms of nacre can be obtained by conventional light-optical methods. For example, we observed super-structures formed by co-oriented nacre platelets as previously identified using X-ray Photo-electron Emission Microscopy (X-PEEM) [Gilbert et al., Journal of the American Chemical Society 2008, 130:17519–17527]. Polarized optical microscopy revealed unprecedented super-structures in the calcitic shell part. This bears, in principle, the potential for in vivo studies, which might be useful for investigating the growth modes of nacre and other shell types. PMID:22967319

Spiritual Coping: A Focus of New Nursing Diagnoses.

PubMed

Cabaço, Sandra Rosado; Caldeira, Sílvia; Vieira, Margarida; Rodgers, Beth

2017-03-01

To define the antecedents, consequents, and attributes of spiritual coping. Rodgers' evolutionary model for concept analysis was used to guide an integrative literature review of qualitative research. Six qualitative articles were included and elements that define and contextualize the concept were identified. Three new nursing diagnoses are proposed, based on qualitative findings. These new diagnoses should be submitted to clinical validation in different cultural and religious backgrounds, but the inclusion in the taxonomy highlights a holistic perspective concerning the spiritual dimension of patients' responses in life and health transitions, and so, bringing the approach to spirituality into nursing practice. Definir os antecedentes, os consequentes e os atributos de coping espiritual. MÉTODOS: Modelo evolucionário de análise de conceitos de Beth Rodgers baseado numa revisão integrativa de literatura de pesquisa qualitativa. Seis pesquisas qualitativas foram incluídas e os elementos que definem e contextualizam o conceito foram identificados. CONCLUSÕES: São propostos três novos diagnósticos de enfermagem, baseados na evidência de estudos qualitativos. IMPLICAÇÕES PARA A PRÁTICA: Estes novos diagnósticos devem ser submetidos a estudos de validação clínica em diferentes contextos culturais e religiosos, e quando incluídos na taxonomia estarão a enfatizar uma perspectiva holística das respostas dos pacientes relacionada à dimensão espiritual e, assim, promovendo a inclusão da espiritualidade na prática clínica. © 2017 NANDA International, Inc.

The evaluation of the interfacial behavior of LaRC-TPI/Graphite Composites

NASA Technical Reports Server (NTRS)

Ogden, A. L.; Wilkes, G. L.; Hyer, M. W.; Loos, A. C.; Muellerleile, J. T.

1992-01-01

Discussed are the results of several approaches recently considered for improving the interfacial adhesion of LaRC-TPI/graphite composites. Two approaches were investigated, namely altering the matrix and altering the fiber. As a result, three types of LaRC-TPI laminates were produced: amorphous/AS-4, amorphous/XAS, and semicrystalline/AS-4. The laminates were characterized using the transverse tensile test, scanning electron microscopy, optical microscopy, and thermal analysis.

In vivo confocal microscopy in dermatology: from research to clinical application

NASA Astrophysics Data System (ADS)

Ulrich, Martina; Lange-Asschenfeldt, Susanne

2013-06-01

Confocal laser scanning microscopy (CLSM) represents an emerging technique for the noninvasive histomorphological analysis of skin in vivo and has shown its applicability for dermatological research as well as its value as an adjunct tool in the clinical management of skin cancer patients. Herein, we aim to give an overview on the current clinical indications for CLSM in dermatology and also highlight the diverse applications of CLSM in dermatological research.

Correlative SEM SERS for quantitative analysis of dimer nanoparticles.

PubMed

Timmermans, F J; Lenferink, A T M; van Wolferen, H A G M; Otto, C

2016-11-14

A Raman microscope integrated with a scanning electron microscope was used to investigate plasmonic structures by correlative SEM-SERS analysis. The integrated Raman-SEM microscope combines high-resolution electron microscopy information with SERS signal enhancement from selected nanostructures with adsorbed Raman reporter molecules. Correlative analysis is performed for dimers of two gold nanospheres. Dimers were selected on the basis of SEM images from multi aggregate samples. The effect of the orientation of the dimer with respect to the polarization state of the laser light and the effect of the particle gap size on the Raman signal intensity is observed. Additionally, calculations are performed to simulate the electric near field enhancement. These simulations are based on the morphologies observed by electron microscopy. In this way the experiments are compared with the enhancement factor calculated with near field simulations and are subsequently used to quantify the SERS enhancement factor. Large differences between experimentally observed and calculated enhancement factors are regularly detected, a phenomenon caused by nanoscale differences between the real and 'simplified' simulated structures. Quantitative SERS experiments reveal the structure induced enhancement factor, ranging from ∼200 to ∼20 000, averaged over the full nanostructure surface. The results demonstrate correlative Raman-SEM microscopy for the quantitative analysis of plasmonic particles and structures, thus enabling a new analytical method in the field of SERS and plasmonics.

The Role of IQGAP1 in Breast Carcinoma

DTIC Science & Technology

2011-10-01

study! of! the! pathogenesis! of! breast! cancer.! These! include! analysis ! of! intracellular! signaling!by!Western!blotting,! determination!of! cell...proliferation!by! sulforhodamine!B! staining,! fluorescence: activated!cell!sorting!(FACS)! analysis ,!stable!cell!line!generation,!production!of!and...transduction!using!retroviral! and!lentiviral!supernatants,! immunocytochemistry!and!confocal! laser!microscopy,! immunohistochemistry,!and! analysis

Image Analysis, Microscopic, and Spectrochemical Study of the PVC Dry Blending Process,

DTIC Science & Technology

The dry blending process used in the production of electrical grade pvc formulations has been studies using a combination of image analysis , microscopic...by image analysis techniques. Optical and scanning electron microscopy were used to assess morphological differences. Spectrochemical techniques were used to indicate chemical changes.

3D Image Analysis of Geomaterials using Confocal Microscopy

NASA Astrophysics Data System (ADS)

Mulukutla, G.; Proussevitch, A.; Sahagian, D.

2009-05-01

Confocal microscopy is one of the most significant advances in optical microscopy of the last century. It is widely used in biological sciences but its application to geomaterials lingers due to a number of technical problems. Potentially the technique can perform non-invasive testing on a laser illuminated sample that fluoresces using a unique optical sectioning capability that rejects out-of-focus light reaching the confocal aperture. Fluorescence in geomaterials is commonly induced using epoxy doped with a fluorochrome that is impregnated into the sample to enable discrimination of various features such as void space or material boundaries. However, for many geomaterials, this method cannot be used because they do not naturally fluoresce and because epoxy cannot be impregnated into inaccessible parts of the sample due to lack of permeability. As a result, the confocal images of most geomaterials that have not been pre-processed with extensive sample preparation techniques are of poor quality and lack the necessary image and edge contrast necessary to apply any commonly used segmentation techniques to conduct any quantitative study of its features such as vesicularity, internal structure, etc. In our present work, we are developing a methodology to conduct a quantitative 3D analysis of images of geomaterials collected using a confocal microscope with minimal amount of prior sample preparation and no addition of fluorescence. Two sample geomaterials, a volcanic melt sample and a crystal chip containing fluid inclusions are used to assess the feasibility of the method. A step-by-step process of image analysis includes application of image filtration to enhance the edges or material interfaces and is based on two segmentation techniques: geodesic active contours and region competition. Both techniques have been applied extensively to the analysis of medical MRI images to segment anatomical structures. Preliminary analysis suggests that there is distortion in the shapes of the segmented vesicles, vapor bubbles, and void spaces due to the optical measurements, so corrective actions are being explored. This will establish a practical and reliable framework for an adaptive 3D image processing technique for the analysis of geomaterials using confocal microscopy.

Yining Zeng | NREL

Science.gov Websites

pretreatment conditions and biological digestion methods, which might not be detected by large-scale ) "Coherent Raman Microscopy Analysis of Plant Cell Walls," Biomass Conversion: Methods and Protocols, Methods in Molecular Biology (2012) "Chemical, Ultrastructural and Supramolecular Analysis

Composition analysis of a polymer electrolyte membrane fuel cell microporous layer using scanning transmission X-ray microscopy and near edge X-ray absorption fine structure analysis

NASA Astrophysics Data System (ADS)

George, Michael G.; Wang, Jian; Banerjee, Rupak; Bazylak, Aimy

2016-03-01

The novel application of scanning transmission X-ray microscopy (STXM) to the microporous layer (MPL) of a polymer electrolyte membrane fuel cell is investigated. A spatially resolved chemical component distribution map is obtained for the MPL of a commercially available SGL 25 BC sample. This is achieved with near edge X-ray absorption fine structure spectroscopic analysis. Prior to analysis the sample is embedded in non-reactive epoxy and ultra-microtomed to a thickness of 100 nm. Polytetrafluoroethylene (PTFE), carbon particle agglomerates, and supporting epoxy resin distributions are identified and reconstructed for a scanning area of 6 μm × 6 μm. It is observed that the spatial distribution of PTFE is strongly correlated to the carbon particle agglomerations. Additionally, agglomerate structures of PTFE are identified, possibly indicating the presence of a unique mesostructure in the MPL. STXM analysis is presented as a useful technique for the investigation of chemical species distributions in the MPL.

Spermatozoa quality assessment: a combined holographic and Raman microscopy approach

NASA Astrophysics Data System (ADS)

De Angelis, Annalisa; Ferrara, Maria A.; Di Caprio, Giuseppe; Managò, Stefano; Sirleto, Luigi; Coppola, Giuseppe; De Luca, Anna Chiara

2015-05-01

Semen analysis is widely used as diagnostic tool for assessing male fertility, controlling and managing the animal reproduction. The most important parameters measured in a semen analysis are the morphology and biochemical alterations. For obtaining such information, non-invasive, label-free and non-destructive techniques have to be used. Digital Holography (DH) combined with Raman Spectroscopy (RS) could represent the perfect candidate for a rapid, non-destructive and high-sensitive morphological and biochemical sperm cell analysis. In this study, DH-RS combined approach is used for a complete analysis of single bovine spermatozoa. High-resolution images of bovine sperm have been obtained by DH microscopy from the reconstruction of a single acquired hologram, highlighting in some cases morphological alterations. Quantitative 3D reconstructions of sperm head, both normal and anomalous, have been studied and an unexpected structure of the post-acrosomal region of the head has been detected. Such anomalies have been also confirmed by Raman imaging analysis, suggesting the protein vibrations as associated Raman marker of the defect.

The importance of transmission electron microscopy analysis of spermatozoa: Diagnostic applications and basic research.

PubMed

Moretti, Elena; Sutera, Gaetano; Collodel, Giulia

2016-06-01

This review is aimed at discussing the role of ultrastructural studies on human spermatozoa and evaluating transmission electron microscopy as a diagnostic tool that can complete andrology protocols. It is clear that morphological sperm defects may explain decreased fertilizing potential and acquire particular value in the field of male infertility. Electron microscopy is the best method to identify systematic or monomorphic and non-systematic or polymorphic sperm defects. The systematic defects are characterized by a particular anomaly that affects the vast majority of spermatozoa in a semen sample, whereas a heterogeneous combination of head and tail defects found in variable percentages are typically non-systematic or polymorphic sperm defects. A correct diagnosis of these specific sperm alterations is important for choosing the male infertility's therapy and for deciding to turn to assisted reproduction techniques. Transmission electron microscopy (TEM) also represents a valuable method to explore the in vitro effects of different compounds (for example drugs with potential spermicidal activity) on the morphology of human spermatozoa. Finally, TEM used in combination with immunohistochemical techniques, integrates structural and functional aspects that provide a wide horizon in the understanding of sperm physiology and pathology. transmission electron microscopy: TEM; World Health Organization: WHO; light microscopy: LM; motile sperm organelle morphology examination: MSOME; intracytoplasmic morphologically selected sperm injection: IMSI; intracytoplasmic sperm injection: ICSI; dysplasia of fibrous sheath: DFS; primary ciliary dyskinesia: PCD; outer dense fibers: ODF; assisted reproduction technologies: ART; scanning electron microscopy: SEM; polyvinylpirrolidone: PVP; tert-butylhydroperoxide: TBHP.

Identification of Foreign Particles in Human Tissues using Raman Microscopy.

PubMed

Campion, Alan; Smith, Kenneth J; Fedulov, Alexey V; Gregory, David; Fan, Yuwei; Godleski, John J

2018-06-12

The precise identification of foreign particles in tissue for patient care and research has been studied using polarized light microscopy, electron microscopy with X-ray analysis, and electron diffraction. The goal of this study was to unambiguously identify particles in tissues using a combina-tion of polarized light microscopy and Raman microscopy, which provides chemical composition and microstructural characterization of complex materials with submicron spatial resolution. We designed a model system of stained and unstained cells that contained birefringent talc particles, and systematically investigated the influence of slide and coverslip materials, laser wavelengths, and mounting media on the Raman spectra ob-tained. Hematoxylin and eosin stained slides did not produce useful results because of fluorescence interference from the stains. Unstained cell samples prepared with standard slides and coverslips produce high quality Raman spectra when excited at 532 nm; the spectra are uniquely as-signed to talc. We also obtain high quality Raman spectra specific for talc in unstained tissue samples (pleural tissue following talc pleurodesis and ovarian tissue following long-term perineal talc exposure). Raman microscopy is sufficiently sensitive and compositionally selective to identify particles as small as one micron in diameter. Among commonly used coverslip mounting media, Cytoseal 60 is recommended; Permount was unacceptable due to intense background interference. Raman spectra have been catalogued for thousands of substances, which suggests that this approach is likely to be successful in identifying other particles of interest in tissues, potentially making Raman microscopy a powerful new tool in pathology.

Automated Analysis of siRNA Screens of Virus Infected Cells Based on Immunofluorescence Microscopy

NASA Astrophysics Data System (ADS)

Matula, Petr; Kumar, Anil; Wörz, Ilka; Harder, Nathalie; Erfle, Holger; Bartenschlager, Ralf; Eils, Roland; Rohr, Karl

We present an image analysis approach as part of a high-throughput microscopy screening system based on cell arrays for the identification of genes involved in Hepatitis C and Dengue virus replication. Our approach comprises: cell nucleus segmentation, quantification of virus replication level in cells, localization of regions with transfected cells, cell classification by infection status, and quality assessment of an experiment. The approach is fully automatic and has been successfully applied to a large number of cell array images from screening experiments. The experimental results show a good agreement with the expected behavior of positive as well as negative controls and encourage the application to screens from further high-throughput experiments.

Direct ultrasonic-assisted synthesis of sphere-like nanocrystals of spinel Co3O4 and Mn3O4.

PubMed

Askarinejad, Azadeh; Morsali, Ali

2009-01-01

A simple sonochemical method was developed to synthesize uniform sphere-like or cubic Co(3)O(4) and Mn(3)O(4) nanocrystals by using acetate salts and sodium hydroxide or tetramethylammonium hydroxide (TMAH) as precursors. Influence of some parameters such as time of reaction, alkali salts, and power of the ultrasound and the molar ratio of the starting materials on the size, morphology and degree of crystallinity of the products was studied. Powder X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), FTIR spectroscopy, Thermal gravimetry analysis and differential thermal analysis (TGA/DTA) were used to characterize the nanocrystals.

Otoconial formation in the fetal rat

NASA Technical Reports Server (NTRS)

Salamat, M. S.; Ross, M. D.; Peacor, D. R.

1980-01-01

Otoconial formation in the fetal rat is examined by scanning and transmission electron microscopy, and by X-ray elemental analysis. The primitive otoconia appear highly organic, but are trigonal in cross section, indicating that they already possess a three-fold axis of symmetry and a complement of calcite. These otoconia develop into spindle-shaped and, subsequently, dumbbell-shaped units. Transmission electron microscopy of dumbbell-shaped otoconia not exposed to fluids during embedment showed that calcite deposits mimicked the arrangement of the organic material. X-ray elemental analysis demonstrated that calcium was present in lower quantities in the central core than peripherally. It is concluded that organic material is essential to otoconial seeding and directs otoconial growth.

Quantitative phase imaging of human red blood cells using phase-shifting white light interference microscopy with colour fringe analysis

NASA Astrophysics Data System (ADS)

Singh Mehta, Dalip; Srivastava, Vishal

2012-11-01

We report quantitative phase imaging of human red blood cells (RBCs) using phase-shifting interference microscopy. Five phase-shifted white light interferograms are recorded using colour charge coupled device camera. White light interferograms were decomposed into red, green, and blue colour components. The phase-shifted interferograms of each colour were then processed by phase-shifting analysis and phase maps for red, green, and blue colours were reconstructed. Wavelength dependent refractive index profiles of RBCs were computed from the single set of white light interferogram. The present technique has great potential for non-invasive determination of refractive index variation and morphological features of cells and tissues.

Scanning electron microscopy fractography analysis of fractured hollow implants.

PubMed

Sbordone, Ludovico; Traini, Tonino; Caputi, Sergio; Scarano, Antonio; Bortolaia, Claudia; Piattelli, Adriano

2010-01-01

Fracture of the implant is one of the possible complications affecting dental implants; it is a rare event but of great clinical relevance. The aim of the present study was to perform a scanning electron microscopy (SEM) fractography evaluation of 7 International Team for oral Implantology (ITI) hollow implants removed because of fracture. The most common clinical risk factors, such as malocclusion, bruxism, and cantilevers on the prosthesis, were absent. Seven fractured ITI hollow implants were retrieved from 5 patients and were analyzed with the use of SEM. SEM analysis showed typical signs of a cleavage-type fracture. Fractures could be due to an association of multiple factors such as fatigue, inner defects, material electrochemical problems, and tensocorrosion.

Fabrication of composite films containing zirconia and cationic polyelectrolytes.

PubMed

Pang, Xin; Zhitomirsky, Igor

2004-03-30

Composite films were prepared by electrophoretic deposition of poly(ethylenimine) or poly(allylamine hydrochloride) combined with cathodic precipitation of zirconia. Films of up to several micrometers thick were obtained on Ni, Pt, stainless-steel, graphite, and carbon-felt substrates. When the concentration of polyelectrolytes in solutions and the deposition time were varied, the amount of the deposited material and its composition can be varied. The electrochemical intercalation of yttria-stabilized zirconia particles into the composite films has been demonstrated. Obtained results pave the way for the electrodeposition of other polymer-ceramic composites. The deposits were studied by thermogravimetric analysis, X-ray diffraction analysis, scanning electron microscopy, and atomic force microscopy. The mechanisms of deposition are discussed.

Quantitative measurements of nanoscale permittivity and conductivity using tuning-fork-based microwave impedance microscopy

NASA Astrophysics Data System (ADS)

Wu, Xiaoyu; Hao, Zhenqi; Wu, Di; Zheng, Lu; Jiang, Zhanzhi; Ganesan, Vishal; Wang, Yayu; Lai, Keji

2018-04-01

We report quantitative measurements of nanoscale permittivity and conductivity using tuning-fork (TF) based microwave impedance microscopy (MIM). The system is operated under the driving amplitude modulation mode, which ensures satisfactory feedback stability on samples with rough surfaces. The demodulated MIM signals on a series of bulk dielectrics are in good agreement with results simulated by finite-element analysis. Using the TF-MIM, we have visualized the evolution of nanoscale conductance on back-gated MoS2 field effect transistors, and the results are consistent with the transport data. Our work suggests that quantitative analysis of mesoscopic electrical properties can be achieved by near-field microwave imaging with small distance modulation.

Study of the Local Horizon. (Spanish Title: Estudio del Horizonte Local.) Estudo do Horizonte Local

NASA Astrophysics Data System (ADS)

Ros, Rosa M.

2009-12-01

The study of the horizon is fundamental to easy the first observations of the students at any education center. A simple model, to be developed in each center, allows to easy the study and comprehension of the rudiments of astronomy. The constructed model is presented in turn as a simple equatorial clock, other models (horizontal and vertical) may be constructed starting from it. El estudio del horizonte es fundamental para poder facilitar las primeras observaciones de los alumnos en un centro educativo. Un simple modelo, que debe realizarse para cada centro, nos permite facilitar el estudio y la comprensión de los primeros rudimentos astronómicos. El modelo construido se presenta a su vez como un sencillo modelo de reloj ecuatorial y a partir de él se pueden construir otros modelos (horizontal y vertical). O estudo do horizonte é fundamental para facilitar as primeiras observações dos alunos num centro educativo. Um modelo simples, que deve ser feito para cada centro, permite facilitar o estudo e a compreensão dos primeiros rudimentos astronômicos. O modelo construído apresenta-se, por sua vez, como um modelo simples de relógio equatorial e a partir dele pode-se construir outros modelos (horizontal e vertical)

A correlação índice espectral vs. luminosidade em QSOs e suas implicações

NASA Astrophysics Data System (ADS)

Garcia-Rissmann, A.

2003-08-01

Estudos de variabilidade de núcleos ativos já demonstraram ser comum o fato de seu contínuo óptico/UV tornar-se mais "duro" à medida que a luminosidade aumenta. Essa tendência ocorre tanto de forma individual quanto global, e pode ter implicações importantes (1) para estudos fotométricos de variabilidade conduzidos numa banda fixa no referencial do observador, comparando objetos a diferentes redshifts, e (2) no cálculo da correção K, com consequente impacto na determinação de massas de buracos negros e bojos de galáxias hospedeiras (através da relação de Magorrian). Confirmo aqui as correlações positivas entre o índice espectral e a luminosidade óptica, utilizando dados espectroscópicos de 11 QSOs monitorados no Brasil e no Chile, durante ~2 anos. O estudo é complementado com parâmetros extraídos de espectros e de dados fotométricos públicos de quasares. Destaco ainda as diferenças observadas em tais correlações para objetos do tipo radio-loud e radio-quiet. Este projeto é financiado pelo I. Milênio/CNPq.

Pigment analysis by Raman microscopy and portable X-ray fluorescence (pXRF) of thirteenth to fourteenth century illuminations and cuttings from Bologna

PubMed Central

Clark, Robin J. H.; Jones, Richard; Gibbs, Robert

2016-01-01

Non-destructive pigment analysis by Raman microscopy (RM) and portable X-ray fluorescence (pXRF) has been carried out on some Bolognese illuminations and cuttings chosen to represent the beginnings, evolution and height of Bolognese illuminated manuscript production. Dating to the thirteenth and fourteenth centuries and held in a private collection, the study provides evidence for the pigments generally used in this period. The results, which are compared with those obtained for other north Italian artwork, show the developments in usage of artistic materials and technique. Also addressed in this study is an examination of the respective roles of RM and pXRF analysis in this area of technical art history. This article is part of the themed issue ‘Raman spectroscopy in art and archaeology’. PMID:27799427

Analysis of Black Bearing Balls from a Space Shuttle Body Flap Actuator

NASA Technical Reports Server (NTRS)

Sovinski, Marjorie F.; Street, Kenneth W.

2005-01-01

A significantly deteriorated ball bearing mechanism from a body flap actuator on Space Shuttle OV-103 was disassembled and the balls submitted for analysis in conjunction with Return to Flight activities. The OV-103 balls, referred to as the "black balls", were subjected to X-ray photoelectron spectroscopy (XPS), Fourier transform infrared (FT-IR) and Raman micro spectroscopy, surface profilometry, and optical and electron microscopy. The spectroscopic results in combination with microscopy analysis allowed a determination of the lubricant degradation pathway. The chemical attack mechanism does not adequately explain the unique visual appearance of the black balls. Numerous efforts have unsuccessfully focused on duplication of the phenomena causing this unique surface structure and appearance of the black balls. Further detail will be presented supporting these conclusions along with plausible explanations of the unique black appearance to the balls.

An Analysis of the Nonlinear Spectral Mixing of Didymium and Soda-Lime Glass Beads Using Hyperspectral Imagery (HSI) Microscopy

DTIC Science & Technology

2014-05-01

2001). Learning with Kernels: Support Vector Machines , Regularization, Optimization, and Beyond. The MIT Press, Cambridge, MA, 644 p. [26] Banerjee...A., Burlina, P., and Broadwater, J., (2007). A Machine Learning Approach for finding hyperspectral endmembers. Proceedings of the IEEE International... lime glass beads using hyperspectral imagery (HSI) microscopy Ronald G. Resmini1*, Robert S. Rand2, David W. Allen3, and Christopher J. Deloye1

Targeted DNA sequencing and in situ mutation analysis using mobile phone microscopy

NASA Astrophysics Data System (ADS)

Kühnemund, Malte; Wei, Qingshan; Darai, Evangelia; Wang, Yingjie; Hernández-Neuta, Iván; Yang, Zhao; Tseng, Derek; Ahlford, Annika; Mathot, Lucy; Sjöblom, Tobias; Ozcan, Aydogan; Nilsson, Mats

2017-01-01

Molecular diagnostics is typically outsourced to well-equipped centralized laboratories, often far from the patient. We developed molecular assays and portable optical imaging designs that permit on-site diagnostics with a cost-effective mobile-phone-based multimodal microscope. We demonstrate that targeted next-generation DNA sequencing reactions and in situ point mutation detection assays in preserved tumour samples can be imaged and analysed using mobile phone microscopy, achieving a new milestone for tele-medicine technologies.

Combination of atomic force microscopy and mass spectrometry for the detection of target protein in the serum samples of children with autism spectrum disorders

NASA Astrophysics Data System (ADS)

Kaysheva, A. L.; Pleshakova, T. O.; Kopylov, A. T.; Shumov, I. D.; Iourov, I. Y.; Vorsanova, S. G.; Yurov, Y. B.; Ziborov, V. S.; Archakov, A. I.; Ivanov, Y. D.

2017-10-01

Possibility of detection of target proteins associated with development of autistic disorders in children with use of combined atomic force microscopy and mass spectrometry (AFM/MS) method is demonstrated. The proposed method is based on the combination of affine enrichment of proteins from biological samples and visualization of these proteins by AFM and MS analysis with quantitative detection of target proteins.

Model-free iterative control of repetitive dynamics for high-speed scanning in atomic force microscopy.

PubMed

Li, Yang; Bechhoefer, John

2009-01-01

We introduce an algorithm for calculating, offline or in real time and with no explicit system characterization, the feedforward input required for repetitive motions of a system. The algorithm is based on the secant method of numerical analysis and gives accurate motion at frequencies limited only by the signal-to-noise ratio and the actuator power and range. We illustrate the secant-solver algorithm on a stage used for atomic force microscopy.

The evaluation of the interfacial behavior of LaRC-TPI/Graphite Composites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ogden, A.L.; Wilkes, G.L.; Hyer, M.W.

1992-07-01

Discussed are the results of several approaches recently considered for improving the interfacial adhesion of LaRC-TPI/graphite composites. Two approaches were investigated, namely altering the matrix and altering the fiber. As a result, three types of LaRC-TPI laminates were produced: amorphous/AS-4, amorphous/XAS, and semicrystalline/AS-4. The laminates were characterized using the transverse tensile test, scanning electron microscopy, optical microscopy, and thermal analysis. 17 refs.

Identification of a Multicomponent Traditional Herbal Medicine by HPLC-MS and Electron and Light Microscopy.

PubMed

Liu, Ju-Han; Cheng, Yung-Yi; Hsieh, Chen-Hsi; Tsai, Tung-Hu

2017-12-15

Commercial pharmaceutical herbal products have enabled people to take traditional Chinese medicine (TCM) in a convenient and accessible form. However, the quantity and quality should be additionally inspected. To address the issue, a combination of chemical and physical inspection methods were developed to evaluate the amount of an herbal formula, Xiang-Sha-Liu-Jun-Zi-Tang (XSLJZT), in clinical TCM practice. A high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS) method with electrospray ionization was developed to measure the herbal biomarkers of guanosine, atractylenolide III, glycyrrhizic acid, dehydrocostus lactone, hesperidin, and oleanolic acid from XSLJZT. Scanning electron microscopy (SEM) photographs and light microscopy photographs with Congo red and iodine-KI staining were used to identify the cellulose fibers and starch content. Furthermore, solubility analysis, swelling power test, and crude fiber analysis were contributed to measure the starch additive in pharmaceutical products. The results demonstrated large variations in the chemical components of different pharmaceutical brands. The SEM photographs revealed that the starch was oval, smooth, and granular, and that the raw herbal powder appears stripy, stretched, and filiform. The stained light microscopy photographs of all of the pharmaceutical products showed added starch and raw herbal powder as extenders. The developed chemical and physical methods provide a standard operating procedure for the quantity control of the herbal pharmaceutical products of XSLJZT.

Diagnostic performance of urine dipstick testing in children with suspected UTI: a systematic review of relationship with age and comparison with microscopy.

PubMed

Mori, R; Yonemoto, N; Fitzgerald, A; Tullus, K; Verrier-Jones, K; Lakhanpaul, M

2010-04-01

Prompt diagnosis of urinary tract infection (UTI) in children is needed to initiate treatment but is difficult to establish without urine testing, and reliance on culture leads to delay. Urine dipsticks are often used as an alternative to microscopy, although the diagnostic performance of dipsticks at different ages has not been established systematically. Studies comparing urine dipstick testing in infants versus older children and urine dipstick versus microscopy were systematically searched and reviewed. Meta-analysis of available studies was conducted. Six studies addressed these questions. The results of meta-analysis showed that the performance of urine dipstick testing was significantly less in the younger children when compared with older children (p < 0.01). Positive likelihood ratio (LR) of both nitrite and leucocyte positive 38.54 [95% confidence interval (CI) 22.49-65.31], negative LR for both negative 0.13 (95% CI 0.07-0.25) are reasonably good, and those for young infants are less reliable [positive LR 7.62 (95% CI 0.95-51.85) and negative LR 0.34 (95% CI 0.66-0.15)]. Comparing microscopy and urine dipstick testing, using bacterial colony count on urine culture showed no significant difference between the two methods. Urine dipstick testing is more effective for diagnosis of UTI in children over 2 years than for younger children.

Investigation of Local Ordering in Amorphous Materials.

NASA Astrophysics Data System (ADS)

Fan, Gary Guoyou

The intent of the research described in this dissertation, as indicated by the title, is to provide a better understanding of the structure of amorphous material. The possibility of using electron microscopy to study the amorphous structure is investigated. Chapter 1 gives a brief introduction to the understanding and modeling of the amorphous structure, electron microscopy and the image analysis in general. The difficulty of using 2-D images to infer 3-D structures information is illustrated in Chapter 2, where it is shown that some high resolution images are not qualitatively different from images of white -noises weak-phase objects or those of random atomic arrangements. The means of obtaining statistical information from these images is given in Chapters 3 and 5, where the quantitative differences between experimental images and simulated white-noise or simulated images corresponding to random arrangements are revealed. The use of image processing techniques in electron microscopy and the possible artifacts are presented in Chapter 4. The pattern recognition technique outlined in Chapter 6 demonstrates a feasible mode of scanning transition electron microscope operation. Statistical analysis can be effectively performed on a large number of nano-diffraction patterns from, for example, locally ordered samples. Some recent developments in physics as well as in electron microscopy are briefly reviewed, and their possible applications in the study of amorphous structures are discussed in Chapter 7.

Evaluation of erythrocyte dysmorphism by light microscopy with lowering of the condenser lens: A simple and efficient method.

PubMed

Barros Silva, Gyl Eanes; Costa, Roberto Silva; Ravinal, Roberto Cuan; Saraiva e Silva, Jucélia; Dantas, Marcio; Coimbra, Terezila Machado

2010-03-01

To demonstrate that the evaluation of erythrocyte dysmorphism by light microscopy with lowering of the condenser lens (LMLC) is useful to identify patients with a haematuria of glomerular or non-glomerular origin. A comparative double-blind study between phase contrast microscopy (PCM) and LMLC is reported to evaluate the efficacy of these techniques. Urine samples of 39 patients followed up for 9 months were analyzed, and classified as glomerular and non-glomerular haematuria. The different microscopic techniques were compared using receiver-operator curve (ROC) analysis and area under curve (AUC). Reproducibility was assessed by coefficient of variation (CV). Specific cut-offs were set for each method according to their best rate of specificity and sensitivity as follows: 30% for phase contrast microscopy and 40% for standard LMLC, reaching in the first method the rate of 95% and 100% of sensitivity and specificity, respectively, and in the second method the rate of 90% and 100% of sensitivity and specificity, respectively. In ROC analysis, AUC for PCM was 0.99 and AUC for LMLC was 0.96. The CV was very similar in glomerular haematuria group for PCM (35%) and LMLC (35.3%). LMLC proved to be effective in contributing to the direction of investigation of haematuria, toward the nephrological or urological side. This method can substitute PCM when this equipment is not available.

Effects of dignity therapy on terminally ill patients: a systematic review.

PubMed

Donato, Suzana Cristina Teixeira; Matuoka, Jéssica Yumi; Yamashita, Camila Cristófero; Salvetti, Marina de Goés

2016-01-01

Analyzing the evidence of the effects of dignity therapy onterminally ill patients. A Systematic review of the literature conducted using the search strategy in six databases. Inclusion criteria were primary studies, excluding literature reviews (systematic or not) and conceptual articles. Ten articles were analyzed regarding method, results and evidence level. Dignity therapy improved the sense of meaning andpurpose, will to live, utility, quality of life, dignity and family appreciationin studies with a higher level of evidence. The effects are not well established in relation to depression, anxiety, spirituality and physical symptoms. Studies with a moderate to high level of evidence have shown increased sense of dignity, will to live and sense of purpose. Further studies should be developed to increase knowledge about dignity therapy. Analisar as evidências sobre os efeitos da terapia da dignidade para pacientes em fase terminal de vida. Revisão sistemática da literatura realizada em seis bases de dados na estratégia de busca. Os critérios de inclusão foram estudos primários, excluindo-se revisões da literatura (sistemáticas ou não) e artigos conceituais. Dez artigos foram analisados quanto ao método, aos resultados e nível de evidência. Nos estudos com maior nível de evidência, a terapia da dignidade melhorou o senso de significado, propósito, vontade de viver, utilidade, qualidade de vida, dignidade e apreciação familiar.Os efeitos não estão bem estabelecidos em relação à depressão, ansiedade, espiritualidade e aos sintomas físicos. Os estudos de nível de evidência de moderado a alto demonstraram aumento do senso de dignidade, vontade de viver e senso de propósito. Mais estudos devem ser desenvolvidos para ampliar o conhecimento sobre a terapia da dignidade.

Intraoperative confocal microscopy in the visualization of 5-aminolevulinic acid fluorescence in low-grade gliomas.

PubMed

Sanai, Nader; Snyder, Laura A; Honea, Norissa J; Coons, Stephen W; Eschbacher, Jennifer M; Smith, Kris A; Spetzler, Robert F

2011-10-01

Greater extent of resection (EOR) for patients with low-grade glioma (LGG) corresponds with improved clinical outcome, yet remains a central challenge to the neurosurgical oncologist. Although 5-aminolevulinic acid (5-ALA)-induced tumor fluorescence is a strategy that can improve EOR in gliomas, only glioblastomas routinely fluoresce following 5-ALA administration. Intraoperative confocal microscopy adapts conventional confocal technology to a handheld probe that provides real-time fluorescent imaging at up to 1000× magnification. The authors report a combined approach in which intraoperative confocal microscopy is used to visualize 5-ALA tumor fluorescence in LGGs during the course of microsurgical resection. Following 5-ALA administration, patients with newly diagnosed LGG underwent microsurgical resection. Intraoperative confocal microscopy was conducted at the following points: 1) initial encounter with the tumor; 2) the midpoint of tumor resection; and 3) the presumed brain-tumor interface. Histopathological analysis of these sites correlated tumor infiltration with intraoperative cellular tumor fluorescence. Ten consecutive patients with WHO Grades I and II gliomas underwent microsurgical resection with 5-ALA and intraoperative confocal microscopy. Macroscopic tumor fluorescence was not evident in any patient. However, in each case, intraoperative confocal microscopy identified tumor fluorescence at a cellular level, a finding that corresponded to tumor infiltration on matched histological analyses. Intraoperative confocal microscopy can visualize cellular 5-ALA-induced tumor fluorescence within LGGs and at the brain-tumor interface. To assess the clinical value of 5-ALA for high-grade gliomas in conjunction with neuronavigation, and for LGGs in combination with intraoperative confocal microscopy and neuronavigation, a Phase IIIa randomized placebo-controlled trial (BALANCE) is underway at the authors' institution.

An update: improvements in imaging perfluorocarbon-mounted plant leaves with implications for studies of plant pathology, physiology, development and cell biology

PubMed Central

Littlejohn, George R.; Mansfield, Jessica C.; Christmas, Jacqueline T.; Witterick, Eleanor; Fricker, Mark D.; Grant, Murray R.; Smirnoff, Nicholas; Everson, Richard M.; Moger, Julian; Love, John

2014-01-01

Plant leaves are optically complex, which makes them difficult to image by light microscopy. Careful sample preparation is therefore required to enable researchers to maximize the information gained from advances in fluorescent protein labeling, cell dyes and innovations in microscope technologies and techniques. We have previously shown that mounting leaves in the non-toxic, non-fluorescent perfluorocarbon (PFC), perfluorodecalin (PFD) enhances the optical properties of the leaf with minimal impact on physiology. Here, we assess the use of the PFCs, PFD, and perfluoroperhydrophenanthrene (PP11) for in vivo plant leaf imaging using four advanced modes of microscopy: laser scanning confocal microscopy (LSCM), two-photon fluorescence microscopy, second harmonic generation microscopy, and stimulated Raman scattering (SRS) microscopy. For every mode of imaging tested, we observed an improved signal when leaves were mounted in PFD or in PP11, compared to mounting the samples in water. Using an image analysis technique based on autocorrelation to quantitatively assess LSCM image deterioration with depth, we show that PP11 outperformed PFD as a mounting medium by enabling the acquisition of clearer images deeper into the tissue. In addition, we show that SRS microscopy can be used to image PFCs directly in the mesophyll and thereby easily delimit the “negative space” within a leaf, which may have important implications for studies of leaf development. Direct comparison of on and off resonance SRS micrographs show that PFCs do not to form intracellular aggregates in live plants. We conclude that the application of PFCs as mounting media substantially increases advanced microscopy image quality of living mesophyll and leaf vascular bundle cells. PMID:24795734

Angular reconstitution-based 3D reconstructions of nanomolecular structures from superresolution light-microscopy images

PubMed Central

Salas, Desirée; Le Gall, Antoine; Fiche, Jean-Bernard; Valeri, Alessandro; Ke, Yonggang; Bron, Patrick; Bellot, Gaetan

2017-01-01

Superresolution light microscopy allows the imaging of labeled supramolecular assemblies at a resolution surpassing the classical diffraction limit. A serious limitation of the superresolution approach is sample heterogeneity and the stochastic character of the labeling procedure. To increase the reproducibility and the resolution of the superresolution results, we apply multivariate statistical analysis methods and 3D reconstruction approaches originally developed for cryogenic electron microscopy of single particles. These methods allow for the reference-free 3D reconstruction of nanomolecular structures from two-dimensional superresolution projection images. Since these 2D projection images all show the structure in high-resolution directions of the optical microscope, the resulting 3D reconstructions have the best possible isotropic resolution in all directions. PMID:28811371

AutoMicromanager: A microscopy scripting toolkit for LABVIEW and other programming environments

NASA Astrophysics Data System (ADS)

Ashcroft, Brian Alan; Oosterkamp, Tjerk

2010-11-01

We present a scripting toolkit for the acquisition and analysis of a wide variety of imaging data by integrating the ease of use of various programming environments such as LABVIEW, IGOR PRO, MATLAB, SCILAB, and others. This toolkit is designed to allow the user to quickly program a variety of standard microscopy components for custom microscopy applications allowing much more flexibility than other packages. Included are both programming tools as well as graphical user interface classes allowing a standard, consistent, and easy to maintain scripting environment. This programming toolkit allows easy access to most commonly used cameras, stages, and shutters through the Micromanager project so the scripter can focus on their custom application instead of boilerplate code generation.

AutoMicromanager: a microscopy scripting toolkit for LABVIEW and other programming environments.

PubMed

Ashcroft, Brian Alan; Oosterkamp, Tjerk

2010-11-01

We present a scripting toolkit for the acquisition and analysis of a wide variety of imaging data by integrating the ease of use of various programming environments such as LABVIEW, IGOR PRO, MATLAB, SCILAB, and others. This toolkit is designed to allow the user to quickly program a variety of standard microscopy components for custom microscopy applications allowing much more flexibility than other packages. Included are both programming tools as well as graphical user interface classes allowing a standard, consistent, and easy to maintain scripting environment. This programming toolkit allows easy access to most commonly used cameras, stages, and shutters through the Micromanager project so the scripter can focus on their custom application instead of boilerplate code generation.

Macromolecular structure of cellulose studied by second-harmonic generation imaging microscopy

NASA Astrophysics Data System (ADS)

Brown, R. Malcom; Millard, Andrew C.; Campagnola, Paul J.

2003-11-01

The macromolecular structure of purified cellulose samples is studied by second-harmonic generation (SHG) imaging microscopy. We show that the SHG contrast in both Valonia and Acetobacter cellulose strongly resembles that of collagen from animal tissues, both in terms of morphology and polarization anisotropy. Polarization analysis shows that microfibrils in each lamella are highly aligned and ordered and change directions by 90° in adjacent lamellae. The angular dependence of the SHG intensity fits well to a cos2 θ distribution, which is characteristic of the electric dipole interaction. Enzymatic degradation of Valonia fibers by cellulase is followed in real time by SHG imaging and results in exponential decay kinetics, showing that SHG imaging microscopy is ideal for monitoring dynamics in biological systems.

A Survey of the Use of Iterative Reconstruction Algorithms in Electron Microscopy

PubMed Central

Otón, J.; Vilas, J. L.; Kazemi, M.; Melero, R.; del Caño, L.; Cuenca, J.; Conesa, P.; Gómez-Blanco, J.; Marabini, R.; Carazo, J. M.

2017-01-01

One of the key steps in Electron Microscopy is the tomographic reconstruction of a three-dimensional (3D) map of the specimen being studied from a set of two-dimensional (2D) projections acquired at the microscope. This tomographic reconstruction may be performed with different reconstruction algorithms that can be grouped into several large families: direct Fourier inversion methods, back-projection methods, Radon methods, or iterative algorithms. In this review, we focus on the latter family of algorithms, explaining the mathematical rationale behind the different algorithms in this family as they have been introduced in the field of Electron Microscopy. We cover their use in Single Particle Analysis (SPA) as well as in Electron Tomography (ET). PMID:29312997

Scanning probe microscopy for the analysis of composite Ti/hydrocarbon plasma polymer thin films

NASA Astrophysics Data System (ADS)

Choukourov, A.; Grinevich, A.; Slavinska, D.; Biederman, H.; Saito, N.; Takai, O.

2008-03-01

Composite Ti/hydrocarbon plasma polymer films with different Ti concentration were deposited on silicon by dc magnetron sputtering of titanium in an atmosphere of argon and hexane. As measured by Kelvin force microscopy and visco-elastic atomic force microscopy, respectively, surface potential and hardness increase with increasing Ti content. Adhesion force to silicon and to fibrinogen molecules was stronger for the Ti-rich films as evaluated from the AFM force-distance curves. Fibrinogen forms a very soft layer on these composites with part of the protein molecules embedded in the outermost region of the plasma polymer. An increase of the surface charge due to fibrinogen adsorption has been observed and attributed to positively charged αC domains of fibrinogen molecule.

Pump-probe imaging of pigmented cutaneous melanoma primary lesions gives insight into metastatic potential

PubMed Central

Robles, Francisco E.; Deb, Sanghamitra; Wilson, Jesse W.; Gainey, Christina S.; Selim, M. Angelica; Mosca, Paul J.; Tyler, Douglas S.; Fischer, Martin C.; Warren, Warren S.

2015-01-01

Metastatic melanoma is associated with a poor prognosis, but no method reliably predicts which melanomas of a given stage will ultimately metastasize and which will not. While sentinel lymph node biopsy (SLNB) has emerged as the most powerful predictor of metastatic disease, the majority of people dying from metastatic melanoma still have a negative SLNB. Here we analyze pump-probe microscopy images of thin biopsy slides of primary melanomas to assess their metastatic potential. Pump-probe microscopy reveals detailed chemical information of melanin with subcellular spatial resolution. Quantification of the molecular signatures without reference standards is achieved using a geometrical representation of principal component analysis. Melanin structure is analyzed in unison with the chemical information by applying principles of mathematical morphology. Results show that melanin in metastatic primary lesions has lower chemical diversity than non-metastatic primary lesions, and contains two distinct phenotypes that are indicative of aggressive disease. Further, the mathematical morphology analysis reveals melanin in metastatic primary lesions has a distinct “dusty” quality. Finally, a statistical analysis shows that the combination of the chemical information with spatial structures predicts metastatic potential with much better sensitivity than SLNB and high specificity, suggesting pump-probe microscopy can be an important tool to help predict the metastatic potential of melanomas. PMID:26417529

Multifrequency spectrum analysis using fully digital G Mode-Kelvin probe force microscopy.

PubMed

Collins, Liam; Belianinov, Alex; Somnath, Suhas; Rodriguez, Brian J; Balke, Nina; Kalinin, Sergei V; Jesse, Stephen

2016-03-11

Since its inception over two decades ago, Kelvin probe force microscopy (KPFM) has become the standard technique for characterizing electrostatic, electrochemical and electronic properties at the nanoscale. In this work, we present a purely digital, software-based approach to KPFM utilizing big data acquisition and analysis methods. General mode (G-Mode) KPFM works by capturing the entire photodetector data stream, typically at the sampling rate limit, followed by subsequent de-noising, analysis and compression of the cantilever response. We demonstrate that the G-Mode approach allows simultaneous multi-harmonic detection, combined with on-the-fly transfer function correction-required for quantitative CPD mapping. The KPFM approach outlined in this work significantly simplifies the technique by avoiding cumbersome instrumentation optimization steps (i.e. lock in parameters, feedback gains etc), while also retaining the flexibility to be implemented on any atomic force microscopy platform. We demonstrate the added advantages of G-Mode KPFM by allowing simultaneous mapping of CPD and capacitance gradient (C') channels as well as increased flexibility in data exploration across frequency, time, space, and noise domains. G-Mode KPFM is particularly suitable for characterizing voltage sensitive materials or for operation in conductive electrolytes, and will be useful for probing electrodynamics in photovoltaics, liquids and ionic conductors.

Nano titania aided clustering and adhesion of beneficial bacteria to plant roots to enhance crop growth and stress management.

PubMed

Palmqvist, N G M; Bejai, S; Meijer, J; Seisenbaeva, G A; Kessler, V G

2015-05-13

A novel use of Titania nanoparticles as agents in the nano interface interaction between a beneficial plant growth promoting bacterium (Bacillus amyloliquefaciens UCMB5113) and oilseed rape plants (Brassica napus) for protection against the fungal pathogen Alternaria brassicae is presented. Two different TiO2 nanoparticle material were produced by the Sol-Gel approach, one using the patented Captigel method and the other one applying TiBALDH precursor. The particles were characterized by transmission electron microscopy, thermogravimetric analysis, X-ray diffraction, dynamic light scattering and nano particle tracking analysis. Scanning electron microscopy showed that the bacterium was living in clusters on the roots and the combined energy-dispersive X-ray spectroscopy analysis revealed that titanium was present in these cluster formations. Confocal laser scanning microscopy further demonstrated an increased bacterial colonization of Arabidopsis thaliana roots and a semi-quantitative microscopic assay confirmed an increased bacterial adhesion to the roots. An increased amount of adhered bacteria was further confirmed by quantitative fluorescence measurements. The degree of infection by the fungus was measured and quantified by real-time-qPCR. Results showed that Titania nanoparticles increased adhesion of beneficial bacteria on to the roots of oilseed rape and protected the plants against infection.

Nano titania aided clustering and adhesion of beneficial bacteria to plant roots to enhance crop growth and stress management

NASA Astrophysics Data System (ADS)

Palmqvist, N. G. M.; Bejai, S.; Meijer, J.; Seisenbaeva, G. A.; Kessler, V. G.

2015-05-01

A novel use of Titania nanoparticles as agents in the nano interface interaction between a beneficial plant growth promoting bacterium (Bacillus amyloliquefaciens UCMB5113) and oilseed rape plants (Brassica napus) for protection against the fungal pathogen Alternaria brassicae is presented. Two different TiO2 nanoparticle material were produced by the Sol-Gel approach, one using the patented Captigel method and the other one applying TiBALDH precursor. The particles were characterized by transmission electron microscopy, thermogravimetric analysis, X-ray diffraction, dynamic light scattering and nano particle tracking analysis. Scanning electron microscopy showed that the bacterium was living in clusters on the roots and the combined energy-dispersive X-ray spectroscopy analysis revealed that titanium was present in these cluster formations. Confocal laser scanning microscopy further demonstrated an increased bacterial colonization of Arabidopsis thaliana roots and a semi-quantitative microscopic assay confirmed an increased bacterial adhesion to the roots. An increased amount of adhered bacteria was further confirmed by quantitative fluorescence measurements. The degree of infection by the fungus was measured and quantified by real-time-qPCR. Results showed that Titania nanoparticles increased adhesion of beneficial bacteria on to the roots of oilseed rape and protected the plants against infection.

Comparison of nitrogen adsorption and transmission electron microscopy analyses for structural characterization of carbon nanotubes

NASA Astrophysics Data System (ADS)

Abbaslou, Reza Malek; Vosoughi, Vahid; Dalai, Ajay K.

2017-10-01

Carbon nanotubes (CNTs) are different from other porous substrates such as activated carbon due to their high external surfaces. This structural feature can lead in some uncertainties in the results of nitrogen adsorption analysis for characterization of CNTs. In this paper, the results of microscopic analyses and nitrogen adsorption method for characterization of carbon nanotubes were compared. Five different types of CNTs with different structures were either synthesized or purchased. The CNT samples were characterized by high resolution transmission electron microscopy (HRTEM), scanning electron microscopy (SEM) and N2 adsorption analysis. The comparisons between the results from the microscopic analyses and N2 adsorption showed that the total pore volume and BET surface measurements include the internal and external porosity of CNTs. Therefore, the interpretation of N2 adsorption data required accurate TEM analysis. In addition, the evaluation of pore size distribution curves from all CNT samples in this study and several instances in the literature revealed the presence of a common peak in the range of 2-5 nm. This peak does not explain the inner pore size distribution. The presence of this common peak can be attributed to the strong adsorption of N2 on the junction of touched and crossed nanotubes.

Label-free in vivo analysis of intracellular lipid droplets in the oleaginous microalga Monoraphidium neglectum by coherent Raman scattering microscopy.

PubMed

Jaeger, Daniel; Pilger, Christian; Hachmeister, Henning; Oberländer, Elina; Wördenweber, Robin; Wichmann, Julian; Mussgnug, Jan H; Huser, Thomas; Kruse, Olaf

2016-10-21

Oleaginous photosynthetic microalgae hold great promise as non-food feedstocks for the sustainable production of bio-commodities. The algal lipid quality can be analysed by Raman micro-spectroscopy, and the lipid content can be imaged in vivo in a label-free and non-destructive manner by coherent anti-Stokes Raman scattering (CARS) microscopy. In this study, both techniques were applied to the oleaginous microalga Monoraphidium neglectum, a biotechnologically promising microalga resistant to commonly applied lipid staining techniques. The lipid-specific CARS signal was successfully separated from the interfering two-photon excited fluorescence of chlorophyll and for the first time, lipid droplet formation during nitrogen starvation could directly be analysed. We found that the neutral lipid content deduced from CARS image analysis strongly correlated with the neutral lipid content measured gravimetrically and furthermore, that the relative degree of unsaturation of fatty acids stored in lipid droplets remained similar. Interestingly, the lipid profile during cellular adaption to nitrogen starvation showed a two-phase characteristic with initially fatty acid recycling and subsequent de novo lipid synthesis. This works demonstrates the potential of quantitative CARS microscopy as a label-free lipid analysis technique for any microalgal species, which is highly relevant for future biotechnological applications and to elucidate the process of microalgal lipid accumulation.

Observations and Modeling of the Green Ocean Amazon 2014/15: Transmission Electron Microscopy Analysis of Aerosol Particles Field Campaign Report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buseck, Peter

2016-03-01

During two Intensive Operational Periods (IOP), we collected samples at 3-hour intervals for transmission electron microscopy analysis. The resulting transmission electron microscopy images and compositions were analyzed for the samples of interest. Further analysis will be done especially for the plume of interest. We found solid spherical organic particles from rebounded samples collected with Professor Scot Martin’s group (Harvard University). Approximately 30% of the rebounded particles at 95% relative humidity were spherical organic particles. Their sources and formation process are not known, but such spherical particles could be solid and will have heterogeneous chemical reactions. We observed many organic particlesmore » that are internally mixed with inorganic elements such as potassium and nitrogen. They are either homogeneously mixed or have inorganic cores with organic aerosol coatings. Samples collected from the Manaus, Brazil, pollution plume included many nano-size soot particles mixed with organic material and sulfate. Aerosol particles from clean periods included organic aerosol particles, sulfate, sea salt, dust, and primary biogenic aerosol particles. There was more dust, primary biogenic aerosol, and tar balls in samples taken during IOP1 than those taken during IOP2. Many dust particles were found between March 2 and 3.« less

Super-Resolution Imaging Strategies for Cell Biologists Using a Spinning Disk Microscope

PubMed Central

Hosny, Neveen A.; Song, Mingying; Connelly, John T.; Ameer-Beg, Simon; Knight, Martin M.; Wheeler, Ann P.

2013-01-01

In this study we use a spinning disk confocal microscope (SD) to generate super-resolution images of multiple cellular features from any plane in the cell. We obtain super-resolution images by using stochastic intensity fluctuations of biological probes, combining Photoactivation Light-Microscopy (PALM)/Stochastic Optical Reconstruction Microscopy (STORM) methodologies. We compared different image analysis algorithms for processing super-resolution data to identify the most suitable for analysis of particular cell structures. SOFI was chosen for X and Y and was able to achieve a resolution of ca. 80 nm; however higher resolution was possible >30 nm, dependant on the super-resolution image analysis algorithm used. Our method uses low laser power and fluorescent probes which are available either commercially or through the scientific community, and therefore it is gentle enough for biological imaging. Through comparative studies with structured illumination microscopy (SIM) and widefield epifluorescence imaging we identified that our methodology was advantageous for imaging cellular structures which are not immediately at the cell-substrate interface, which include the nuclear architecture and mitochondria. We have shown that it was possible to obtain two coloured images, which highlights the potential this technique has for high-content screening, imaging of multiple epitopes and live cell imaging. PMID:24130668

Second harmonic generation microscopy analysis of extracellular matrix changes in human idiopathic pulmonary fibrosis

PubMed Central

Tilbury, Karissa; Hocker, James; Wen, Bruce L.; Sandbo, Nathan; Singh, Vikas; Campagnola, Paul J.

2014-01-01

Abstract. Patients with idiopathic fibrosis (IPF) have poor long-term survival as there are limited diagnostic/prognostic tools or successful therapies. Remodeling of the extracellular matrix (ECM) has been implicated in IPF progression; however, the structural consequences on the collagen architecture have not received considerable attention. Here, we demonstrate that second harmonic generation (SHG) and multiphoton fluorescence microscopy can quantitatively differentiate normal and IPF human tissues. For SHG analysis, we developed a classifier based on wavelet transforms, principle component analysis, and a K-nearest-neighbor algorithm to classify the specific alterations of the collagen structure observed in IPF tissues. The resulting ROC curves obtained by varying the numbers of principal components and nearest neighbors yielded accuracies of >95%. In contrast, simpler metrics based on SHG intensity and collagen coverage in the image provided little or no discrimination. We also characterized the change in the elastin/collagen balance by simultaneously measuring the elastin autofluorescence and SHG intensities and found that the IPF tissues were less elastic relative to collagen. This is consistent with known mechanical consequences of the disease. Understanding ECM remodeling in IPF via nonlinear optical microscopy may enhance our ability to differentiate patients with rapid and slow progression and, thus, provide better prognostic information. PMID:25134793

Nano titania aided clustering and adhesion of beneficial bacteria to plant roots to enhance crop growth and stress management

PubMed Central

Palmqvist, N. G. M.; Bejai, S.; Meijer, J.; Seisenbaeva, G. A.; Kessler, V. G.

2015-01-01

A novel use of Titania nanoparticles as agents in the nano interface interaction between a beneficial plant growth promoting bacterium (Bacillus amyloliquefaciens UCMB5113) and oilseed rape plants (Brassica napus) for protection against the fungal pathogen Alternaria brassicae is presented. Two different TiO2 nanoparticle material were produced by the Sol-Gel approach, one using the patented Captigel method and the other one applying TiBALDH precursor. The particles were characterized by transmission electron microscopy, thermogravimetric analysis, X-ray diffraction, dynamic light scattering and nano particle tracking analysis. Scanning electron microscopy showed that the bacterium was living in clusters on the roots and the combined energy-dispersive X-ray spectroscopy analysis revealed that titanium was present in these cluster formations. Confocal laser scanning microscopy further demonstrated an increased bacterial colonization of Arabidopsis thaliana roots and a semi-quantitative microscopic assay confirmed an increased bacterial adhesion to the roots. An increased amount of adhered bacteria was further confirmed by quantitative fluorescence measurements. The degree of infection by the fungus was measured and quantified by real-time-qPCR. Results showed that Titania nanoparticles increased adhesion of beneficial bacteria on to the roots of oilseed rape and protected the plants against infection. PMID:25970693